Gut microbiome composition is altered in Parkinson’s disease (PD), the fastest-growing neurological condition, that is characterized by neurodegeneration, motor dysfunction, and is frequently accompanied by gastrointestinal (GI) symptoms. Notably, microbial taxa with anti-inflammatory properties are consistently depleted in PD patients compared to controls. To explore whether specific gut bacteria may be disease-protective, we assembled a microbial consortium of 8 human-associated taxa that are reduced in individuals with PD. Treatment of α-synuclein overexpressing (Thy1-ASO) mice, an animal model of PD, with this consortium improved motor and GI deficits. A single bacterial species from this consortium, Faecalibacterium prausnitzii, was sufficient to correct gut microbiome deviations in Thy1-ASO mice, induce anti-inflammatory immune responses, and promote protective colonic gene expression profiles. Accordingly, oral treatment with F. prausnitzii robustly ameliorated motor and GI deficits and reduced α-synuclein aggregates in the brain. These findings support the emerging hypothesis of functional contributions by the microbiome to PD outcomes, and embolden the development of potential probiotic therapies to treat motor and non-motor symptoms.
", "doi": "10.1038/s41531-026-01287-x", "pmcid": "PMC13077053", "issn": "2373-8057", "publisher": "Nature Publishing Group", "publication": "npj Parkinson's Disease", "publication_date": "2026-04-13", "series_number": "1", "volume": "12", "issue": "1", "pages": "94" }, { "id": "authors:r8hcr-3n621", "collection": "authors", "collection_id": "r8hcr-3n621", "cite_using_url": "https://authors.library.caltech.edu/records/r8hcr-3n621", "type": "article", "title": "Faecalibacterium prausnitzii, depleted in the Parkinson's disease microbiome, improves motor deficits in \u03b1-synuclein overexpressing mice", "author": [ { "family_name": "Moiseyenko", "given_name": "Anastasiya", "clpid": "Moiseyenko-Anastasiya" }, { "family_name": "Antonello", "given_name": "Giacomo" }, { "family_name": "Schonhoff", "given_name": "Aubrey M.", "orcid": "0000-0002-1801-1125", "clpid": "Schonhoff-Aubrey-M" }, { "family_name": "Boktor", "given_name": "Joseph C.", "orcid": "0000-0003-2456-1913", "clpid": "Boktor-Joseph-C" }, { "family_name": "Long", "given_name": "Kaelyn" }, { "family_name": "Dirks", "given_name": "Blake" }, { "family_name": "Oguienko", "given_name": "Anastasiya D.", "clpid": "Oguienko-Anastasiya-D" }, { "family_name": "Winnett", "given_name": "Alexander Viloria", "orcid": "0000-0002-7338-5605", "clpid": "Winnett-Alexander-Viloria" }, { "family_name": "Simpson", "given_name": "Patrick", "clpid": "Simpson-Patrick" }, { "family_name": "Daeizadeh", "given_name": "Dorsa" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Krajmalnik-Brown", "given_name": "Rosa" }, { "family_name": "Segata", "given_name": "Nicola" }, { "family_name": "Waldron", "given_name": "Levi D." }, { "family_name": "Mazmanian", "given_name": "Sarkis K.", "orcid": "0000-0003-2713-1513", "clpid": "Mazmanian-S-K" } ], "abstract": "Gut microbiome composition is altered in Parkinson’s disease (PD), the fastest-growing neurological condition, that is characterized by neurodegeneration, motor dysfunction, and is frequently accompanied by gastrointestinal (GI) symptoms. Notably, microbial taxa with anti-inflammatory properties are consistently depleted in PD patients compared to controls. To explore whether specific gut bacteria may be disease-protective, we assembled a microbial consortium of 8 human-associated taxa that are reduced in individuals with PD. Treatment of α-synuclein overexpressing (Thy1-ASO) mice, an animal model of PD, with this consortium improved motor and GI deficits. A single bacterial species from this consortium, Faecalibacterium prausnitzii, was sufficient to correct gut microbiome deviations in Thy1-ASO mice, induce anti-inflammatory immune responses, and promote protective colonic gene expression profiles. Accordingly, oral treatment with F. prausnitzii robustly ameliorated motor and GI deficits and reduced α-synuclein aggregates in the brain. These findings support the emerging hypothesis of functional contributions by the microbiome to PD outcomes, and embolden the development of potential probiotic therapies to treat motor and non-motor symptoms.
", "doi": "10.1038/s41531-026-01287-x", "issn": "2373-8057", "publisher": "Nature Publishing Group", "publication": "npj Parkinson's Disease", "publication_date": "2026-03-05" }, { "id": "authors:jxwan-q7e32", "collection": "authors", "collection_id": "jxwan-q7e32", "cite_using_url": "https://authors.library.caltech.edu/records/jxwan-q7e32", "type": "article", "title": "Inducible, but not constitutive, pancreatic REG/Reg isoforms are regulated by intestinal microbiota and pancreatic diseases", "author": [ { "family_name": "Zhou", "given_name": "Yixuan D.", "orcid": "0000-0002-5522-0671" }, { "family_name": "Komnick", "given_name": "Macy R.", "orcid": "0000-0001-8201-6310" }, { "family_name": "Sepulveda", "given_name": "Fabiola", "orcid": "0009-0007-9219-9828" }, { "family_name": "Liu", "given_name": "Grace", "orcid": "0000-0002-1758-4415" }, { "family_name": "Nieves-Ortiz", "given_name": "Elida", "orcid": "0009-0004-6655-6025" }, { "family_name": "Meador", "given_name": "Kelsey", "orcid": "0009-0005-0734-4839" }, { "family_name": "Ndatabaye", "given_name": "Ornella" }, { "family_name": "Fatkhullina", "given_name": "Aliia" }, { "family_name": "Bozicevich", "given_name": "Asha" }, { "family_name": "Juengel", "given_name": "Braden" }, { "family_name": "Wu-Woods", "given_name": "Natalie J.", "orcid": "0000-0001-5070-9091", "clpid": "Wu-Woods-Natalie-J" }, { "family_name": "Naydenkov", "given_name": "Paulina M.", "clpid": "Naydenkov-Paulina-M" }, { "family_name": "Kent", "given_name": "Johnathan", "orcid": "0000-0002-8262-1497" }, { "family_name": "Christiansen", "given_name": "Nathaniel" }, { "family_name": "Madariaga", "given_name": "Maria Lucia", "orcid": "0000-0002-2950-7230" }, { "family_name": "Witkowski", "given_name": "Piotr" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Esterh\u00e1zy", "given_name": "Daria", "orcid": "0000-0002-2427-9873" } ], "abstract": "The REG/Reg gene locus encodes a conserved family of potent antimicrobial but also pancreatitis-associated proteins. Here we investigated whether REG/Reg family members differ in their baseline expression levels and abilities to be regulated in the pancreas and gut upon perturbations. We found, in humans and mice, the pancreas and gut differed in REG/Reg isoform levels and preferences, with the duodenum most resembling the pancreas. Pancreatic acinar cells and intestinal enterocytes were the dominant REG producers. Intestinal symbiotic microbes regulated the expression of the same, select Reg members in gut and pancreas. These Reg members had the most STAT3-binding sites close to the transcription start sites and were partially IL-22 dependent. We thus categorized them as “inducible” and others as “constitutive”. Indeed, in pancreatic ductal adenocarcinoma and pancreatitis models, only inducible Reg members were upregulated in the pancreas. While intestinal Reg expression remained unchanged upon pancreatic perturbation, pancreatitis altered the microbial composition of the duodenum and feces shortly after disease onset. Our study reveals differential usage and regulation of REG/Reg isoforms as a mechanism for tissue-specific innate immunity, highlights the intimate connection of pancreas and duodenum, and implies a gut-to-pancreas communication axis resulting in a coordinated Reg response.
", "doi": "10.1016/j.mucimm.2025.05.003", "issn": "1933-0219", "publisher": "Elsevier", "publication": "Mucosal Immunology", "publication_date": "2025-08", "series_number": "4", "volume": "18", "issue": "4", "pages": "918\u2013936" }, { "id": "authors:daweg-ss366", "collection": "authors", "collection_id": "daweg-ss366", "cite_using_url": "https://authors.library.caltech.edu/records/daweg-ss366", "type": "article", "title": "COVID-19 study quantifying daily viral loads confirms throat samples are key to early diagnosis", "author": [ { "family_name": "Viloria Winnett", "given_name": "Alexander" }, { "family_name": "Shelby", "given_name": "Natasha", "orcid": "0000-0001-9097-3663", "clpid": "Shelby-Natasha" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "doi": "10.1016/j.diagmicrobio.2025.116742", "issn": "0732-8893", "publisher": "Elsevier", "publication": "Diagnostic Microbiology and Infectious Disease", "publication_date": "2025-04", "series_number": "4", "volume": "111", "issue": "4", "pages": "116742" }, { "id": "authors:rjrx5-9bm15", "collection": "authors", "collection_id": "rjrx5-9bm15", "cite_using_url": "https://authors.library.caltech.edu/records/rjrx5-9bm15", "type": "article", "title": "Microbial-enrichment method enables high-throughput metagenomic characterization from host-rich samples", "author": [ { "family_name": "Wu-Woods", "given_name": "Natalie J.", "orcid": "0000-0001-5070-9091", "clpid": "Wu-Woods-Natalie-J" }, { "family_name": "Barlow", "given_name": "Jacob T.", "orcid": "0000-0002-1842-4835", "clpid": "Barlow-Jacob-T" }, { "family_name": "Trigodet", "given_name": "Florian", "orcid": "0000-0002-4933-2896", "clpid": "Trigodet-Florian" }, { "family_name": "Shaw", "given_name": "Dustin G.", "orcid": "0000-0003-3961-7588", "clpid": "Shaw-Dustin-G" }, { "family_name": "Romano", "given_name": "Anna E.", "orcid": "0000-0002-7148-0668", "clpid": "Romano-Anna-E" }, { "family_name": "Jabri", "given_name": "Bana", "orcid": "0000-0001-7427-4424", "clpid": "Jabri-Bana" }, { "family_name": "Eren", "given_name": "A. Murat", "orcid": "0000-0001-9013-4827" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Host\u2013microbe interactions have been linked to health and disease states through the use of microbial taxonomic profiling, mostly via 16S ribosomal RNA gene sequencing. However, many mechanistic insights remain elusive, in part because studying the genomes of microbes associated with mammalian tissue is difficult due to the high ratio of host to microbial DNA in such samples. Here we describe a microbial-enrichment method (MEM), which we demonstrate on a wide range of sample types, including saliva, stool, intestinal scrapings, and intestinal mucosal biopsies. MEM enabled high-throughput characterization of microbial metagenomes from human intestinal biopsies by reducing host DNA more than 1,000-fold with minimal microbial community changes (roughly 90% of taxa had no significant differences between MEM-treated and untreated control groups). Shotgun sequencing of MEM-treated human intestinal biopsies enabled characterization of both high- and low-abundance microbial taxa, pathways and genes longitudinally along the gastrointestinal tract. We report the construction of metagenome-assembled genomes directly from human intestinal biopsies for bacteria and archaea at relative abundances as low as 1%. Analysis of metagenome-assembled genomes reveals distinct subpopulation structures between the small and large intestine for some taxa. MEM opens a path for the microbiome field to acquire deeper insights into host\u2013microbe interactions by enabling in-depth characterization of host-tissue-associated microbial communities.
", "doi": "10.1038/s41592-023-02025-4", "pmcid": "PMC10885704", "issn": "1548-7091", "publisher": "Nature Publishing Group", "publication": "Nature Methods", "publication_date": "2023-11", "series_number": "11", "volume": "20", "issue": "11", "pages": "1672-1682" }, { "id": "authors:7j7m9-fec46", "collection": "authors", "collection_id": "7j7m9-fec46", "cite_using_url": "https://authors.library.caltech.edu/records/7j7m9-fec46", "type": "article", "title": "Index cases first identified by nasal-swab rapid COVID-19 tests had more transmission to household contacts than cases identified by other test types", "author": [ { "family_name": "Ji", "given_name": "Jenny", "orcid": "0000-0002-7901-5605", "clpid": "Ji-Jenny" }, { "family_name": "Viloria Winnett", "given_name": "Alexander", "orcid": "0000-0002-7338-5605", "clpid": "Winnett-Alexander-Viloria" }, { "family_name": "Shelby", "given_name": "Natasha", "orcid": "0000-0001-9097-3663", "clpid": "Shelby-Natasha" }, { "family_name": "Reyes", "given_name": "Jessica A.", "orcid": "0000-0002-5507-7633", "clpid": "Reyes-Jessica-A" }, { "family_name": "Schlenker", "given_name": "Noah W.", "orcid": "0000-0002-8581-4403", "clpid": "Schlenker-Noah-W" }, { "family_name": "Davich", "given_name": "Hannah", "orcid": "0000-0001-6880-3086", "clpid": "Davich-Hannah" }, { "family_name": "Caldera", "given_name": "Saharai", "orcid": "0000-0001-5057-9186", "clpid": "Caldera-Saharai" }, { "family_name": "Tognazzini", "given_name": "Colten", "orcid": "0000-0002-2754-3588", "clpid": "Tognazzini-Colten" }, { "family_name": "Goh", "given_name": "Ying-Ying", "orcid": "0000-0001-5136-7214", "clpid": "Goh-Ying-Ying" }, { "family_name": "Feaster", "given_name": "Matt", "orcid": "0000-0001-9966-2845", "clpid": "Feaster-Matthew" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "At-home rapid COVID-19 tests in the U.S. utilize nasal-swab specimens and require high viral loads to reliably give positive results. Longitudinal studies from the onset of infection have found infectious virus can present in oral specimens days before nasal. Detection and initiation of infection-control practices may therefore be delayed when nasal-swab rapid tests are used, resulting in greater transmission to contacts. We assessed whether index cases first identified by rapid nasal-swab COVID-19 tests had more transmission to household contacts than index cases who used other test types (tests with higher analytical sensitivity and/or non-nasal specimen types). In this observational cohort study, 370 individuals from 85 households with a recent COVID-19 case were screened at least daily by RT-qPCR on one or more self-collected upper-respiratory specimen types. A two-level random intercept model was used to assess the association between the infection outcome of household contacts and each covariable (household size, race/ethnicity, age, vaccination status, viral variant, infection-control practices, and whether a rapid nasal-swab test was used to initially identify the household index case). Transmission was quantified by adjusted secondary attack rates (aSAR) and adjusted odds ratios (aOR). An aSAR of 53.6% (95% CI 38.8\u201368.3%) was observed among households where the index case first tested positive by a rapid nasal-swab COVID-19 test, which was significantly higher than the aSAR for households where the index case utilized another test type (27.2% 95% CI 19.5\u201335.0%, P = 0.003 pairwise comparisons of predictive margins). We observed an aOR of 4.90 (95% CI 1.65\u201314.56) for transmission to household contacts when a nasal-swab rapid test was used to identify the index case, compared to other test types. Use of nasal-swab rapid COVID-19 tests for initial detection of infection and initiation of infection control may be less effective at limiting transmission to household contacts than other test types.", "doi": "10.1371/journal.pone.0292389", "pmcid": "PMC10553276", "issn": "1932-6203", "publisher": "Public Library of Science", "publication": "PLOS ONE", "publication_date": "2023-10", "series_number": "10", "volume": "18", "issue": "10", "pages": "e0292389" }, { "id": "authors:4rjkx-r3m91", "collection": "authors", "collection_id": "4rjkx-r3m91", "cite_using_url": "https://authors.library.caltech.edu/records/4rjkx-r3m91", "type": "article", "title": "Quantitative whole-tissue 3D imaging reveals bacteria in close association with mouse jejunum mucosa", "author": [ { "family_name": "Poceviciute", "given_name": "Roberta", "orcid": "0000-0002-6649-2170", "clpid": "Poceviciute-Roberta" }, { "family_name": "Bogatyrev", "given_name": "Said R.", "orcid": "0000-0003-0486-9451", "clpid": "Bogatyrev-Said-R" }, { "family_name": "Romano", "given_name": "Anna E.", "orcid": "0000-0002-7148-0668", "clpid": "Romano-Anna-E" }, { "family_name": "Dilmore", "given_name": "Amanda H.", "orcid": "0000-0001-6493-7116", "clpid": "Dilmore-Amanda-H" }, { "family_name": "Mondrag\u00f3n-Palomino", "given_name": "Octavio", "orcid": "0000-0003-1129-4932", "clpid": "Mondrag\u00f3n-Palomino-Octavio" }, { "family_name": "Takko", "given_name": "Heli", "orcid": "0000-0003-2544-409X", "clpid": "Takko-Heli" }, { "family_name": "Pradhan", "given_name": "Ojas", "orcid": "0000-0002-5678-4081", "clpid": "Pradhan-Ojas" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Because the small intestine (SI) epithelium lacks a thick protective mucus layer, microbes that colonize the thin SI mucosa may exert a substantial effect on the host. For example, bacterial colonization of the human SI may contribute to environmental enteropathy dysfunction (EED) in malnourished children. Thus far, potential bacterial colonization of the mucosal surface of the SI has only been documented in disease states, suggesting mucosal colonization is rare, likely requiring multiple perturbations. Furthermore, conclusive proof of bacterial colonization of the SI mucosal surface is challenging, and the three-dimensional (3D) spatial structure of mucosal colonies remains unknown. Here, we tested whether we could induce dense bacterial association with jejunum mucosa by subjecting mice to a combination of malnutrition and oral co-gavage with a bacterial cocktail (E. coli and Bacteroides spp.) known to induce EED. To visualize these events, we optimized our previously developed whole-tissue 3D imaging tools with third-generation hybridization chain reaction (HCR v3.0) probes. Only in mice that were malnourished and gavaged with the bacterial cocktail did we detect dense bacterial clusters surrounding intestinal villi suggestive of colonization. Furthermore, in these mice we detected villus loss, which may represent one possible consequence that bacterial colonization of the SI mucosa has on the host. Our results suggest that dense bacterial colonization of jejunum mucosa is possible in the presence of multiple perturbations and that whole-tissue 3D imaging tools can enable the study of these rare events.
", "doi": "10.1038/s41522-023-00423-2", "pmcid": "PMC10485000", "issn": "2055-5008", "publisher": "Nature Publishing Group", "publication": "npj Biofilms and Microbiomes", "publication_date": "2023-09-07", "volume": "9", "pages": "64" }, { "id": "authors:k79gc-vkp02", "collection": "authors", "collection_id": "k79gc-vkp02", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20230628-344482000.1", "type": "article", "title": "Daily SARS-CoV-2 Nasal Antigen Tests Miss Infected and Presumably Infectious People Due to Viral Load Differences among Specimen Types", "author": [ { "family_name": "Winnett", "given_name": "Alexander Viloria", "clpid": "Winnett-Alexander-Viloria" }, { "family_name": "Akana", "given_name": "Reid", "clpid": "Akana-Reid" }, { "family_name": "Shelby", "given_name": "Natasha", "clpid": "Shelby-Natasha" }, { "family_name": "Davich", "given_name": "Hannah", "clpid": "Davich-Hannah" }, { "family_name": "Caldera", "given_name": "Saharai", "clpid": "Caldera-Saharai" }, { "family_name": "Yamada", "given_name": "Taikun", "clpid": "Yamada-Taikun" }, { "family_name": "Reyna", "given_name": "John Raymond B.", "clpid": "Reyna-John-Raymond-B" }, { "family_name": "Romano", "given_name": "Anna E.", "clpid": "Romano-Anne-E" }, { "family_name": "Carter", "given_name": "Alyssa M.", "clpid": "Carter-Alyssa-M" }, { "family_name": "Kim", "given_name": "Mi Kyung", "clpid": "Kim-Mi-Kyung" }, { "family_name": "Thomson", "given_name": "Matt", "orcid": "0000-0003-1021-1234", "clpid": "Thomson-M-W" }, { "family_name": "Tognazzini", "given_name": "Colten", "clpid": "Tognazzini-Colten" }, { "family_name": "Feaster", "given_name": "Matthew", "clpid": "Feaster-Matthew" }, { "family_name": "Goh", "given_name": "Ying-Ying", "clpid": "Goh-Ying-Ying" }, { "family_name": "Chew", "given_name": "Yap Ching", "clpid": "Chew-Yap-Ching" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "In a recent household transmission study of SARS-CoV-2, we found extreme differences in SARS-CoV-2 viral loads among paired saliva, anterior nares swab (ANS), and oropharyngeal swab specimens collected from the same time point. We hypothesized these differences may hinder low-analytical-sensitivity assays (including antigen rapid diagnostic tests [Ag-RDTs]) by using a single specimen type (e.g., ANS) from reliably detecting infected and infectious individuals. We evaluated daily at-home ANS Ag-RDTs (Quidel QuickVue) in a cross-sectional analysis of 228 individuals and a longitudinal analysis (throughout infection) of 17 individuals enrolled early in the course of infection. Ag-RDT results were compared to reverse transcription-quantitative PCR (RT-qPCR) results and high, presumably infectious viral loads (in each, or any, specimen type). The ANS Ag-RDT correctly detected only 44% of time points from infected individuals on cross-sectional analysis, and this population had an inferred limit of detection of 7.6\u2009\u00d7\u200910\u2076 copies/mL. From the longitudinal cohort, daily Ag-RDT clinical sensitivity was very low (<3%) during the early, preinfectious period of the infection. Further, the Ag-RDT detected \u226463% of presumably infectious time points. The poor observed clinical sensitivity of the Ag-RDT was similar to what was predicted based on quantitative ANS viral loads and the inferred limit of detection of the ANS Ag-RDT being evaluated, indicating high-quality self-sampling. Nasal Ag-RDTs, even when used daily, can miss individuals infected with the Omicron variant and even those presumably infectious. Evaluations of Ag-RDTs for detection of infected or infectious individuals should be compared with a composite (multispecimen) infection status to correctly assess performance.\n\nImportance:\nWe reveal three findings from a longitudinal study of daily nasal antigen rapid diagnostic test (Ag-RDT) evaluated against SARS-CoV-2 viral load quantification in three specimen types (saliva, nasal swab, and throat swab) in participants enrolled at the incidence of infection. First, the evaluated Ag-RDT showed low (44%) clinical sensitivity for detecting infected persons at all infection stages. Second, the Ag-RDT poorly detected (\u226463%) time points that participants had high and presumably infectious viral loads in at least one specimen type. This poor clinical sensitivity to detect infectious individuals is inconsistent with the commonly held view that daily Ag-RDTs have near-perfect detection of infectious individuals. Third, use of a combination nasal-throat specimen type was inferred by viral loads to significantly improve Ag-RDT performance to detect infectious individuals.", "doi": "10.1128/spectrum.01295-23", "pmcid": "PMC10434058", "issn": "2165-0497", "publisher": "American Society for Microbiology", "publication": "Microbiology Spectrum", "publication_date": "2023-07", "series_number": "4", "volume": "11", "issue": "4", "pages": "e01295-23" }, { "id": "authors:8xx2v-fk653", "collection": "authors", "collection_id": "8xx2v-fk653", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20230323-759607000.4", "type": "article", "title": "Extreme differences in SARS-CoV-2 viral loads among respiratory specimen types during presumed pre-infectious and infectious periods", "author": [ { "family_name": "Winnett", "given_name": "Alexander Viloria", "orcid": "0000-0002-7338-5605", "clpid": "Winnett-Alexander-Viloria" }, { "family_name": "Akana", "given_name": "Reid", "orcid": "0000-0003-4422-587X", "clpid": "Akana-Reid" }, { "family_name": "Shelby", "given_name": "Natasha", "orcid": "0000-0001-9097-3663", "clpid": "Shelby-Natasha" }, { "family_name": "Davich", "given_name": "Hannah", "orcid": "0000-0001-6880-3086", "clpid": "Davich-Hannah" }, { "family_name": "Caldera", "given_name": "Saharai", "orcid": "0000-0001-5057-9186", "clpid": "Caldera-Saharai" }, { "family_name": "Yamada", "given_name": "Taikun", "clpid": "Yamada-Taikun" }, { "family_name": "Reyna", "given_name": "John Raymond B.", "clpid": "Reyna-John-Raymond-B" }, { "family_name": "Romano", "given_name": "Anna E.", "orcid": "0000-0002-7148-0668", "clpid": "Romano-Anne-E" }, { "family_name": "Carter", "given_name": "Alyssa M.", "orcid": "0000-0002-2776-9421", "clpid": "Carter-Alyssa-M" }, { "family_name": "Kim", "given_name": "Mi Kyung", "clpid": "Kim-Mi-Kyung" }, { "family_name": "Thomson", "given_name": "Matt", "orcid": "0000-0003-1021-1234", "clpid": "Thomson-M-W" }, { "family_name": "Tognazzini", "given_name": "Colten", "orcid": "0000-0002-2754-3588", "clpid": "Tognazzini-Colten" }, { "family_name": "Feaster", "given_name": "Matthew", "orcid": "0000-0001-9966-2845", "clpid": "Feaster-Matthew" }, { "family_name": "Goh", "given_name": "Ying-Ying", "orcid": "0000-0001-5136-7214", "clpid": "Goh-Ying-Ying" }, { "family_name": "Chew", "given_name": "Yap Ching", "orcid": "0000-0002-1686-6541", "clpid": "Chew-Yap-Ching" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "SARS-CoV-2 viral-load measurements from a single-specimen type are used to establish diagnostic strategies, interpret clinical-trial results for vaccines and therapeutics, model viral transmission, and understand virus\u2013host interactions. However, measurements from a single-specimen type are implicitly assumed to be representative of other specimen types. We quantified viral-load timecourses from individuals who began daily self-sampling of saliva, anterior-nares (nasal), and oropharyngeal (throat) swabs before or at the incidence of infection with the Omicron variant. Viral loads in different specimen types from the same person at the same timepoint exhibited extreme differences, up to 109 copies/mL. These differences were not due to variation in sample self-collection, which was consistent. For most individuals, longitudinal viral-load timecourses in different specimen types did not correlate. Throat-swab and saliva viral loads began to rise as many as 7 days earlier than nasal-swab viral loads in most individuals, leading to very low clinical sensitivity of nasal swabs during the first days of infection. Individuals frequently exhibited presumably infectious viral loads in one specimen type while viral loads were low or undetectable in other specimen types. Therefore, defining an individual as infectious based on assessment of a single-specimen type underestimates the infectious period, and overestimates the ability of that specimen type to detect infectious individuals. For diagnostic COVID-19 testing, these three single-specimen types have low clinical sensitivity, whereas a combined throat\u2013nasal swab, and assays with high analytical sensitivity, was inferred to have significantly better clinical sensitivity to detect presumed pre-infectious and infectious individuals.", "doi": "10.1093/pnasnexus/pgad033", "pmcid": "PMC10013338", "issn": "2752-6542", "publisher": "Oxford University Press", "publication": "PNAS Nexus", "publication_date": "2023-03", "series_number": "3", "volume": "2", "issue": "3", "pages": "pgad033" }, { "id": "authors:0mjqz-64458", "collection": "authors", "collection_id": "0mjqz-64458", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20230130-263514000.2", "type": "article", "title": "Laboratory Evaluation Links Some False-Positive COVID-19 Antigen Test Results Observed in a Field Study to a Specific Lot of Test Strips", "author": [ { "family_name": "Carter", "given_name": "Alyssa M.", "orcid": "0000-0002-2776-9421", "clpid": "Carter-Alyssa-M" }, { "family_name": "Winnett", "given_name": "Alexander Viloria", "orcid": "0000-0002-7338-5605", "clpid": "Winnett-Alexander-Viloria" }, { "family_name": "Romano", "given_name": "Anna E.", "orcid": "0000-0002-7148-0668", "clpid": "Romano-Anne-E" }, { "family_name": "Akana", "given_name": "Reid", "orcid": "0000-0003-4422-587X", "clpid": "Akana-Reid" }, { "family_name": "Shelby", "given_name": "Natasha", "orcid": "0000-0001-9097-3663", "clpid": "Shelby-Natasha" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "During a household-transmission field study using COVID-19 antigen rapid diagnostic tests (Ag-RDT), a common test strip lot was identified among three participants with false-positive results. In blinded laboratory evaluation, this lot, exhibited a significantly higher false-positive rate than other lots. Because a positive Ag-RDT result often prompts action, reducing lot-specific false positives can maintain confidence and actionability of true-positive Ag-RDT results.", "doi": "10.1093/ofid/ofac701", "pmcid": "PMC9887260", "issn": "2328-8957", "publisher": "Oxford University Press", "publication": "Open Forum Infectious Diseases", "publication_date": "2023-01-30", "series_number": "1", "volume": "10", "issue": "1", "pages": "Art. No. ofac701" }, { "id": "authors:sh6x3-qaj93", "collection": "authors", "collection_id": "sh6x3-qaj93", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20230130-262818000.1", "type": "article", "title": "GATA4 controls regionalization of tissue immunity and commensal-driven immunopathology", "author": [ { "family_name": "Earley", "given_name": "Zachary M.", "orcid": "0000-0001-9748-7262", "clpid": "Earley-Zachary-M" }, { "family_name": "Lisicka", "given_name": "Wioletta", "orcid": "0000-0003-2623-5185", "clpid": "Lisicka-Wioletta" }, { "family_name": "Sifakis", "given_name": "Joseph J.", "clpid": "Sifakis-Joseph-J" }, { "family_name": "Aguirre-Gamboa", "given_name": "Ra\u00fal", "orcid": "0000-0003-2505-3574", "clpid": "Aguirre-Gamboa-Ra\u00fal" }, { "family_name": "Kowalczyk", "given_name": "Anita", "orcid": "0000-0002-4217-8857", "clpid": "Kowalczyk-Anita" }, { "family_name": "Barlow", "given_name": "Jacob T.", "orcid": "0000-0002-1842-4835", "clpid": "Barlow-Jacob-T" }, { "family_name": "Shaw", "given_name": "Dustin G.", "orcid": "0000-0003-3961-7588", "clpid": "Shaw-Dustin-G" }, { "family_name": "Discepolo", "given_name": "Valentina", "orcid": "0000-0001-6158-0545", "clpid": "Discepolo-Valentina" }, { "family_name": "Tan", "given_name": "Ineke L.", "orcid": "0000-0001-8159-2367", "clpid": "Tan-Ineke-L" }, { "family_name": "Gona", "given_name": "Saideep", "orcid": "0000-0002-7469-2607", "clpid": "Gona-Saideep" }, { "family_name": "Ernest", "given_name": "Jordan D.", "orcid": "0000-0003-0857-217X", "clpid": "Ernest-Jordan-B" }, { "family_name": "Matzinger", "given_name": "Polly", "clpid": "Matzinger-Polly" }, { "family_name": "Barreiro", "given_name": "Luis B.", "orcid": "0000-0001-9456-367X", "clpid": "Barreiro-Luis-B" }, { "family_name": "Morgun", "given_name": "Andrey", "orcid": "0000-0002-8924-6108", "clpid": "Morgun-Andrey" }, { "family_name": "Bendelac", "given_name": "Albert", "orcid": "0000-0003-1859-206X", "clpid": "Bendelac-Albert" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Shulzhenko", "given_name": "Natalia", "orcid": "0000-0003-0077-8456", "clpid": "Shulzhenko-Natalia" }, { "family_name": "Riesenfeld", "given_name": "Samantha J.", "orcid": "0000-0001-9144-021X", "clpid": "Riesenfeld-Samantha-J" }, { "family_name": "Jabri", "given_name": "Bana", "clpid": "Jabri-Bana" } ], "abstract": "There is growing recognition that regionalization of bacterial colonization and immunity along the intestinal tract has an important role in health and disease. Yet, the mechanisms underlying intestinal regionalization and its dysregulation in disease are not well understood. This study found that regional epithelial expression of the transcription factor GATA4 controls bacterial colonization and inflammatory tissue immunity in the proximal small intestine by regulating retinol metabolism and luminal IgA. Furthermore, in mice without jejunal GATA4 expression, the commensal segmented filamentous bacteria promoted pathogenic inflammatory immune responses that disrupted barrier function and increased mortality upon Citrobacter rodentium infection. In celiac disease patients, low GATA4 expression was associated with metabolic alterations, mucosal Actinobacillus, and increased IL-17 immunity. Taken together, these results reveal broad impacts of GATA4-regulated intestinal regionalization on bacterial colonization and tissue immunity, highlighting an elaborate interdependence of intestinal metabolism, immunity, and microbiota in homeostasis and disease.", "doi": "10.1016/j.immuni.2022.12.009", "issn": "1074-7613", "publisher": "Cell Press", "publication": "Immunity", "publication_date": "2023-01-10", "series_number": "1", "volume": "56", "issue": "1", "pages": "43-57.e10" }, { "id": "authors:00s15-xar08", "collection": "authors", "collection_id": "00s15-xar08", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20221110-430693700.10", "type": "article", "title": "Morning SARS-CoV-2 Testing Yields Better Detection of Infection Due to Higher Viral Loads in Saliva and Nasal Swabs upon Waking", "author": [ { "family_name": "Winnett", "given_name": "Alexander Viloria", "orcid": "0000-0002-7338-5605", "clpid": "Winnett-Alexander-Viloria" }, { "family_name": "Porter", "given_name": "Michael K.", "orcid": "0000-0002-0777-7563", "clpid": "Porter-Michael-K" }, { "family_name": "Romano", "given_name": "Anna E.", "orcid": "0000-0002-7148-0668", "clpid": "Romano-Anne-E" }, { "family_name": "Savela", "given_name": "Emily S.", "orcid": "0000-0001-9614-4276", "clpid": "Savela-Emily-S" }, { "family_name": "Akana", "given_name": "Reid", "orcid": "0000-0003-4422-587X", "clpid": "Akana-Reid" }, { "family_name": "Shelby", "given_name": "Natasha", "orcid": "0000-0001-9097-3663", "clpid": "Shelby-Natasha" }, { "family_name": "Reyes", "given_name": "Jessica A.", "orcid": "0000-0002-5507-7633", "clpid": "Reyes-Jessica-A" }, { "family_name": "Schlenker", "given_name": "Noah W.", "orcid": "0000-0002-8581-4403", "clpid": "Schlenker-Noah-W" }, { "family_name": "Cooper", "given_name": "Matthew M.", "orcid": "0000-0002-5868-5159", "clpid": "Cooper-Matthew-M" }, { "family_name": "Carter", "given_name": "Alyssa M.", "orcid": "0000-0002-2776-9421", "clpid": "Carter-Alyssa-M" }, { "family_name": "Ji", "given_name": "Jenny", "orcid": "0000-0002-7901-5605", "clpid": "Ji-Jenny" }, { "family_name": "Barlow", "given_name": "Jacob T.", "orcid": "0000-0002-1842-4835", "clpid": "Barlow-Jacob-T" }, { "family_name": "Tognazzini", "given_name": "Colten", "orcid": "0000-0002-2754-3588", "clpid": "Tognazzini-Colten" }, { "family_name": "Feaster", "given_name": "Matthew", "orcid": "0000-0001-9966-2845", "clpid": "Feaster-Matthew" }, { "family_name": "Goh", "given_name": "Ying-Ying", "orcid": "0000-0001-5136-7214", "clpid": "Goh-Ying-Ying" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Optimizing specimen collection methods to achieve the most reliable SARS-CoV-2 detection for a given diagnostic sensitivity would improve testing and minimize COVID-19 outbreaks. From September 2020 to April 2021, we performed a household-transmission study in which participants self-collected specimens every morning and evening throughout acute SARS-CoV-2 infection. Seventy mildly symptomatic participants collected saliva, and of those, 29 also collected nasal swab specimens. Viral load was quantified in 1,194 saliva and 661 nasal swab specimens using a high-analytical-sensitivity reverse transcription-quantitative PCR (RT-qPCR) assay. Viral loads in both saliva and nasal swab specimens were significantly higher in morning-collected specimens than in evening-collected specimens after symptom onset. This aspect of the biology of SARS-CoV-2 infection has implications for diagnostic testing. We infer that morning collection would have resulted in significantly improved detection and that this advantage would be most pronounced for tests with low to moderate analytical sensitivity. Collecting specimens for COVID-19 testing in the morning offers a simple and low-cost improvement to clinical diagnostic sensitivity of low- to moderate-analytical-sensitivity tests.\n\nImportance: Our findings suggest that collecting saliva and nasal swab specimens in the morning immediately after waking yields higher SARS-CoV-2 viral loads than collection later in the day. The higher viral loads from morning specimen collection are predicted to significantly improve detection of SARS-CoV-2 in symptomatic individuals, particularly when using moderate- to low-analytical-sensitivity COVID-19 diagnostic tests, such as rapid antigen tests.", "doi": "10.1128/spectrum.03873-22", "pmcid": "PMC9769854", "issn": "2165-0497", "publisher": "American Society for Microbiology", "publication": "Microbiology Spectrum", "publication_date": "2022-12-21", "series_number": "6", "volume": "10", "issue": "6", "pages": "e03873-22" }, { "id": "authors:f4njc-jvc50", "collection": "authors", "collection_id": "f4njc-jvc50", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20220428-203848852", "type": "article", "title": "Three-dimensional imaging for the quantification of spatial patterns in microbiota of the intestinal mucosa", "author": [ { "family_name": "Mondrag\u00f3n-Palomino", "given_name": "Octavio", "orcid": "0000-0003-1129-4932", "clpid": "Mondrag\u00f3n-Palomino-Octavio" }, { "family_name": "Poceviciute", "given_name": "Roberta", "orcid": "0000-0002-6649-2170", "clpid": "Poceviciute-Roberta" }, { "family_name": "Lignell", "given_name": "Antti", "orcid": "0000-0001-7664-5583", "clpid": "Lignell-Antti" }, { "family_name": "Griffiths", "given_name": "Jessica A.", "orcid": "0000-0002-5586-1567", "clpid": "Griffiths-Jessica-A" }, { "family_name": "Takko", "given_name": "Heli", "orcid": "0000-0003-2544-409X", "clpid": "Takko-Heli" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Improving our understanding of host\u2013microbe relationships in the gut requires the ability to both visualize and quantify the spatial organization of microbial communities in their native orientation with the host tissue. We developed a systematic procedure to quantify the three-dimensional (3D) spatial structure of the native mucosal microbiota in any part of the intestines with taxonomic and high spatial resolution. We performed a 3D biogeographical analysis of the microbiota of mouse cecal crypts at different stages of antibiotic exposure. By tracking eubacteria and four dominant bacterial taxa, we found that the colonization of crypts by native bacteria is a dynamic and spatially organized process. Ciprofloxacin treatment drastically reduced bacterial loads and eliminated Muribaculaceae (or all Bacteroidetes entirely) even 10 d after recovery when overall bacterial loads returned to preantibiotic levels. Our 3D quantitative imaging approach revealed that the bacterial colonization of crypts is organized in a spatial pattern that consists of clusters of adjacent colonized crypts that are surrounded by unoccupied crypts, and that this spatial pattern is resistant to the elimination of Muribaculaceae or of all Bacteroidetes by ciprofloxacin. Our approach also revealed that the composition of cecal crypt communities is diverse and that Lactobacilli were found closer to the lumen than Bacteroidetes, Ruminococcaceae, and Lachnospiraceae, regardless of antibiotic exposure. Finally, we found that crypts communities with similar taxonomic composition were physically closer to each other than communities that were taxonomically different.", "doi": "10.1073/pnas.2118483119", "pmcid": "PMC9171773", "issn": "0027-8424", "publisher": "National Academy of Sciences", "publication": "Proceedings of the National Academy of Sciences", "publication_date": "2022-05-03", "series_number": "18", "volume": "119", "issue": "18", "pages": "Art. No. e2118483119" }, { "id": "authors:r8qh8-y4065", "collection": "authors", "collection_id": "r8qh8-y4065", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20210407-080559241", "type": "article", "title": "Quantitative SARS-CoV-2 Viral-Load Curves in Paired Saliva Samples and Nasal Swabs Inform Appropriate Respiratory Sampling Site and Analytical Test Sensitivity Required for Earliest Viral Detection", "author": [ { "family_name": "Savela", "given_name": "Emily S.", "orcid": "0000-0001-9614-4276", "clpid": "Savela-Emily-S" }, { "family_name": "Winnett", "given_name": "Alexander Viloria", "orcid": "0000-0002-7338-5605", "clpid": "Winnett-Alexander-Viloria" }, { "family_name": "Romano", "given_name": "Anna E.", "orcid": "0000-0003-1871-1727", "clpid": "Romano-Anna-E" }, { "family_name": "Porter", "given_name": "Michael K.", "orcid": "0000-0002-0777-7563", "clpid": "Porter-Michael-K" }, { "family_name": "Shelby", "given_name": "Natasha", "orcid": "0000-0001-9097-3663", "clpid": "Shelby-Natasha" }, { "family_name": "Akana", "given_name": "Reid", "orcid": "0000-0003-4422-587X", "clpid": "Akana-Reid" }, { "family_name": "Ji", "given_name": "Jenny", "orcid": "0000-0002-7901-5605", "clpid": "Ji-Jenny" }, { "family_name": "Cooper", "given_name": "Matthew M.", "orcid": "0000-0002-5868-5159", "clpid": "Cooper-Matthew-M" }, { "family_name": "Schlenker", "given_name": "Noah W.", "orcid": "0000-0002-8581-4403", "clpid": "Schlenker-Noah-W" }, { "family_name": "Reyes", "given_name": "Jessica A.", "orcid": "0000-0002-5507-7633", "clpid": "Reyes-Jessica-A" }, { "family_name": "Carter", "given_name": "Alyssa M.", "orcid": "0000-0002-2776-9421", "clpid": "Carter-Alyssa-M" }, { "family_name": "Barlow", "given_name": "Jacob T.", "orcid": "0000-0002-1842-4835", "clpid": "Barlow-Jacob-T" }, { "family_name": "Tognazzini", "given_name": "Colten", "clpid": "Tognazzini-Colten" }, { "family_name": "Feaster", "given_name": "Matthew", "orcid": "0000-0001-9966-2845", "clpid": "Feaster-Matthew" }, { "family_name": "Goh", "given_name": "Ying-Ying", "orcid": "0000-0001-5136-7214", "clpid": "Goh-Ying-Ying" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Early detection of SARS-CoV-2 infection is critical to reduce asymptomatic and presymptomatic transmission, curb the spread of variants, and maximize treatment efficacy. Low-analytical-sensitivity nasal-swab testing is commonly used for surveillance and symptomatic testing, but the ability of these tests to detect the earliest stages of infection has not been established. In this study, conducted between September 2020 and June 2021 in the greater Los Angeles County, California, area, initially SARS-CoV-2-negative household contacts of individuals diagnosed with COVID-19 prospectively self-collected paired anterior-nares nasal-swab and saliva samples twice daily for viral-load quantification by high-sensitivity reverse-transcription quantitative PCR (RT-qPCR) and digital-RT-PCR assays. We captured viral-load profiles from the incidence of infection for seven individuals and compared diagnostic sensitivities between respiratory sites. Among unvaccinated persons, testing saliva with a high-analytical-sensitivity assay detected infection up to 4.5\u2009days before viral loads in nasal swabs reached concentrations detectable by low-analytical-sensitivity nasal-swab tests. For most participants, nasal swabs reached higher peak viral loads than saliva but were undetectable or at lower loads during the first few days of infection. High-analytical-sensitivity saliva testing was most reliable for earliest detection. Our study illustrates the value of acquiring early (within hours after a negative high-sensitivity test) viral-load profiles to guide the appropriate analytical sensitivity and respiratory site for detecting earliest infections. Such data are challenging to acquire but critical to designing optimal testing strategies with emerging variants in the current pandemic and to respond to future viral pandemics.", "doi": "10.1128/JCM.01785-21", "pmcid": "PMC8849374", "issn": "0095-1137", "publisher": "American Society for Microbiology", "publication": "Journal of Clinical Microbiology", "publication_date": "2022-02", "series_number": "2", "volume": "60", "issue": "2", "pages": "Art. No. e01785-21" }, { "id": "authors:nepdr-6d707", "collection": "authors", "collection_id": "nepdr-6d707", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20200813-130113364", "type": "article", "title": "Single-cell measurement of higher-order 3D genome organization with scSPRITE", "author": [ { "family_name": "Arrastia", "given_name": "Mary V.", "orcid": "0000-0002-0723-3574", "clpid": "Arrastia-Mary-V" }, { "family_name": "Jachowicz", "given_name": "Joanna W.", "orcid": "0000-0002-1599-682X", "clpid": "Jachowicz-Joanna-W" }, { "family_name": "Ollikainen", "given_name": "Noah", "orcid": "0000-0002-1174-2400", "clpid": "Ollikainen-Noah" }, { "family_name": "Curtis", "given_name": "Matthew S.", "orcid": "0000-0002-9662-3266", "clpid": "Curtis-Matthew-S" }, { "family_name": "Lai", "given_name": "Charlotte", "clpid": "Lai-Charlotte" }, { "family_name": "Quinodoz", "given_name": "Sofia A.", "orcid": "0000-0003-1862-5204", "clpid": "Quinodoz-Sofia-A" }, { "family_name": "Selck", "given_name": "David A.", "orcid": "0000-0002-0591-4165", "clpid": "Selck-David-A" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Guttman", "given_name": "Mitchell", "orcid": "0000-0003-4748-9352", "clpid": "Guttman-M" } ], "abstract": "Although three-dimensional (3D) genome organization is central to many aspects of nuclear function, it has been difficult to measure at the single-cell level. To address this, we developed 'single-cell split-pool recognition of interactions by tag extension' (scSPRITE). scSPRITE uses split-and-pool barcoding to tag DNA fragments in the same nucleus and their 3D spatial arrangement. Because scSPRITE measures multiway DNA contacts, it generates higher-resolution maps within an individual cell than can be achieved by proximity ligation. We applied scSPRITE to thousands of mouse embryonic stem cells and detected known genome structures, including chromosome territories, active and inactive compartments, and topologically associating domains (TADs) as well as long-range inter-chromosomal structures organized around various nuclear bodies. We observe that these structures exhibit different levels of heterogeneity across the population, with TADs representing dynamic units of genome organization across cells. We expect that scSPRITE will be a critical tool for studying genome structure within heterogeneous populations.", "doi": "10.1038/s41587-021-00998-1", "issn": "1087-0156", "publisher": "Nature Publishing Group", "publication": "Nature Biotechnology", "publication_date": "2022-01", "series_number": "1", "volume": "40", "issue": "1", "pages": "64-73" }, { "id": "authors:tmw0j-mph48", "collection": "authors", "collection_id": "tmw0j-mph48", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20211110-202208524", "type": "article", "title": "Quantitative sequencing clarifies the role of disruptor taxa, oral microbiota, and strict anaerobes in the human small-intestine microbiome", "author": [ { "family_name": "Barlow", "given_name": "Jacob T.", "orcid": "0000-0002-1842-4835", "clpid": "Barlow-Jacob-T" }, { "family_name": "Leite", "given_name": "Gabriela", "orcid": "0000-0001-5772-7735", "clpid": "Leite-Gabriela" }, { "family_name": "Romano", "given_name": "Anna E.", "orcid": "0000-0003-1871-1727", "clpid": "Romano-Anna-E" }, { "family_name": "Sedighi", "given_name": "Rashin", "clpid": "Sedighi-Rashin" }, { "family_name": "Chang", "given_name": "Christine", "orcid": "0000-0003-0179-7800", "clpid": "Chang-Christine" }, { "family_name": "Celly", "given_name": "Shreya", "orcid": "0000-0002-5896-457X", "clpid": "Celly-Shreya" }, { "family_name": "Rezaie", "given_name": "Ali", "orcid": "0000-0002-0106-372X", "clpid": "Rezaie-Ali" }, { "family_name": "Mathur", "given_name": "Ruchi", "orcid": "0000-0003-1053-6557", "clpid": "Mathur-Ruchi" }, { "family_name": "Pimentel", "given_name": "Mark", "orcid": "0000-0002-0619-5115", "clpid": "Pimentel-Mark" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Background: Upper gastrointestinal (GI) disorders and abdominal pain afflict between 12 and 30% of the worldwide population and research suggests these conditions are linked to the gut microbiome. Although large-intestine microbiota have been linked to several GI diseases, the microbiota of the human small intestine and its relation to human disease has been understudied. The small intestine is the major site for immune surveillance in the gut, and compared with the large intestine, it has greater than 100 times the surface area and a thinner and more permeable mucus layer. \n\nResults: Using quantitative sequencing, we evaluated total and taxon-specific absolute microbial loads from 250 duodenal-aspirate samples and 21 paired duodenum-saliva samples from participants in the REIMAGINE study. Log-transformed total microbial loads spanned 5 logs and were normally distributed. Paired saliva-duodenum samples suggested potential transmission of oral microbes to the duodenum, including organisms from the HACEK group. Several taxa, including Klebsiella, Escherichia, Enterococcus, and Clostridium, seemed to displace strict anaerobes common in the duodenum, so we refer to these taxa as disruptors. Disruptor taxa were enriched in samples with high total microbial loads and in individuals with small intestinal bacterial overgrowth (SIBO). Absolute loads of disruptors were associated with more severe GI symptoms, highlighting the value of absolute taxon quantification when studying small-intestine health and function. \n\nConclusion: This study provides the largest dataset of the absolute abundance of microbiota from the human duodenum to date. The results reveal a clear relationship between the oral microbiota and the duodenal microbiota and suggest an association between the absolute abundance of disruptor taxa, SIBO, and the prevalence of severe GI symptoms.", "doi": "10.1186/s40168-021-01162-2", "pmcid": "PMC8561862", "issn": "2049-2618", "publisher": "Springer", "publication": "Microbiome", "publication_date": "2021-11-02", "volume": "9", "pages": "Art. No. 214" }, { "id": "authors:6ez4q-hx548", "collection": "authors", "collection_id": "6ez4q-hx548", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20220512-142847077", "type": "article", "title": "Modulating Factors of Household Transmission of SARS\u2011CoV\u20112 in a Community-Based Study", "author": [ { "family_name": "Ji", "given_name": "Jenny", "orcid": "0000-0002-7901-5605", "clpid": "Ji-Jenny" }, { "family_name": "Winnett", "given_name": "Alexander", "orcid": "0000-0002-7338-5605", "clpid": "Winnett-Alexander" }, { "family_name": "Shelby", "given_name": "Natasha", "orcid": "0000-0001-9097-3663", "clpid": "Shelby-Natasha" }, { "family_name": "Reyes", "given_name": "Jessica A.", "orcid": "0000-0002-5507-7633", "clpid": "Reyes-Jessica-A" }, { "family_name": "Akana", "given_name": "Reid", "orcid": "0000-0003-4422-587X", "clpid": "Akana-Reid" }, { "family_name": "Schlenker", "given_name": "Noah W.", "orcid": "0000-0002-8581-4403", "clpid": "Schlenker-Noah-W" }, { "family_name": "Savela", "given_name": "Emily S.", "orcid": "0000-0001-9614-4276", "clpid": "Savela-Emily-S" }, { "family_name": "Romano", "given_name": "Anna", "orcid": "0000-0003-1871-1727", "clpid": "Romano-Anna-E" }, { "family_name": "Porter", "given_name": "Michael K.", "orcid": "0000-0002-0777-7563", "clpid": "Porter-Michael-K" }, { "family_name": "Carter", "given_name": "Alyssa M.", "orcid": "0000-0002-2776-9421", "clpid": "Carter-Alyssa-M" }, { "family_name": "Cooper", "given_name": "Matthew M.", "orcid": "0000-0002-5868-5159", "clpid": "Cooper-Matthew-M" }, { "family_name": "Barlow", "given_name": "Jacob T.", "orcid": "0000-0002-1842-4835", "clpid": "Barlow-Jacob-T" }, { "family_name": "Tognazzini", "given_name": "Colten", "clpid": "Tognazzini-Colten" }, { "family_name": "Feaster", "given_name": "Matthew", "orcid": "0000-0001-9966-2845", "clpid": "Feaster-Matthew" }, { "family_name": "Goh", "given_name": "Ying-Ying", "orcid": "0000-0001-5136-7214", "clpid": "Goh-Ying-Ying" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "[no abstract]", "doi": "10.4269/ajtmh.abstract2021", "issn": "0002-9637", "publisher": "American Society of Tropical Medicine and Hygiene", "publication": "American Journal of Tropical Medicine and Hygiene", "publication_date": "2021-11", "series_number": "5", "volume": "105", "issue": "5", "pages": "313" }, { "id": "authors:en9tt-2np72", "collection": "authors", "collection_id": "en9tt-2np72", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20210908-171123816", "type": "article", "title": "Alterations in the gut microbiota contribute to cognitive impairment induced by the ketogenic diet and hypoxia", "author": [ { "family_name": "Olson", "given_name": "Christine A.", "orcid": "0000-0002-5382-1393", "clpid": "Olson-Christine-A" }, { "family_name": "I\u00f1iguez", "given_name": "Alonso J.", "orcid": "0000-0002-4869-3278", "clpid": "I\u00f1iguez-Alonso-J" }, { "family_name": "Yang", "given_name": "Grace E.", "orcid": "0000-0001-8346-908X", "clpid": "Yang-Grace-E" }, { "family_name": "Fang", "given_name": "Ping", "orcid": "0000-0003-4379-3787", "clpid": "Fang-Ping" }, { "family_name": "Pronovost", "given_name": "Geoffrey N.", "clpid": "Pronovost-Geoffrey-N" }, { "family_name": "Jameson", "given_name": "Kelly G.", "orcid": "0000-0002-9972-9138", "clpid": "Jameson-Kelly-G" }, { "family_name": "Rendon", "given_name": "Tomiko K.", "clpid": "Rendon-Tomiko-K" }, { "family_name": "Paramo", "given_name": "Jorge", "clpid": "Paramo-Jorge" }, { "family_name": "Barlow", "given_name": "Jacob T.", "orcid": "0000-0002-1842-4835", "clpid": "Barlow-Jacob-T" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Hsiao", "given_name": "Elaine Y.", "orcid": "0000-0002-1633-588X", "clpid": "Hsiao-Elaine-Y-Bio" } ], "abstract": "Many genetic and environmental factors increase susceptibility to cognitive impairment (CI), and the gut microbiome is increasingly implicated. However, the identity of gut microbes associated with CI risk, their effects on CI, and their mechanisms remain unclear. Here, we show that a carbohydrate-restricted (ketogenic) diet potentiates CI induced by intermittent hypoxia in mice and alters the gut microbiota. Depleting the microbiome reduces CI, whereas transplantation of the risk-associated microbiome or monocolonization with Bilophila wadsworthia confers CI in mice fed a standard diet. B. wadsworthia and the risk-associated microbiome disrupt hippocampal synaptic plasticity, neurogenesis, and gene expression. The CI is associated with microbiome-dependent increases in intestinal interferon-gamma (IFNg)-producing Th1 cells. Inhibiting Th1 cell development abrogates the adverse effects of both B. wadsworthia and environmental risk factors on CI. Together, these findings identify select gut bacteria that contribute to environmental risk for CI in mice by promoting inflammation and hippocampal dysfunction.", "doi": "10.1016/j.chom.2021.07.004", "pmcid": "PMC8429275", "issn": "1931-3128", "publisher": "Cell Press", "publication": "Cell Host and Microbe", "publication_date": "2021-09-08", "series_number": "9", "volume": "29", "issue": "9", "pages": "1378-1392" }, { "id": "authors:3mzpv-c6728", "collection": "authors", "collection_id": "3mzpv-c6728", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20210706-202344110", "type": "article", "title": "Microbiota regulate social behaviour via stress response neurons in the brain", "author": [ { "family_name": "Wu", "given_name": "Wei-Li", "orcid": "0000-0003-2610-1881", "clpid": "Wu-Wei-Li" }, { "family_name": "Adame", "given_name": "Mark D.", "clpid": "Adame-Mark-D" }, { "family_name": "Liou", "given_name": "Chia-Wei", "orcid": "0000-0002-9003-4065", "clpid": "Liou-Chia-Wei" }, { "family_name": "Barlow", "given_name": "Jacob T.", "orcid": "0000-0002-1842-4835", "clpid": "Barlow-Jacob-T" }, { "family_name": "Lai", "given_name": "Tzu-Ting", "clpid": "Lai-Tzu-Ting" }, { "family_name": "Sharon", "given_name": "Gil", "orcid": "0000-0002-4605-9943", "clpid": "Sharon-Gil" }, { "family_name": "Schretter", "given_name": "Catherine E.", "orcid": "0000-0002-3957-6838", "clpid": "Schretter-Catherine-E" }, { "family_name": "Needham", "given_name": "Brittany D.", "orcid": "0000-0002-0280-1886", "clpid": "Needham-Brittany-D" }, { "family_name": "Wang", "given_name": "Madelyn I.", "orcid": "0000-0001-7576-1179", "clpid": "Wang-Madelyn-I" }, { "family_name": "Tang", "given_name": "Weiyi", "orcid": "0000-0002-1279-1001", "clpid": "Tang-Weiyi" }, { "family_name": "Ousey", "given_name": "James", "orcid": "0000-0003-4886-0053", "clpid": "Ousey-James" }, { "family_name": "Lin", "given_name": "Yuan-Yuan", "clpid": "Lin-Yuan-Yuan" }, { "family_name": "Yao", "given_name": "Tzu-Hsuan", "clpid": "Yao-Tzu-Hsuan" }, { "family_name": "Abdel-Haq", "given_name": "Reem", "orcid": "0000-0002-7418-5736", "clpid": "Abdel-Haq-Reem" }, { "family_name": "Beadle", "given_name": "Keith", "orcid": "0000-0002-5695-6461", "clpid": "Beadle-Keith" }, { "family_name": "Gradinaru", "given_name": "Viviana", "orcid": "0000-0001-5868-348X", "clpid": "Gradinaru-V" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Mazmanian", "given_name": "Sarkis K.", "orcid": "0000-0003-2713-1513", "clpid": "Mazmanian-S-K" } ], "abstract": "Social interactions among animals mediate essential behaviours, including mating, nurturing, and defence. The gut microbiota contribute to social activity in mice, but the gut\u2013brain connections that regulate this complex behaviour and its underlying neural basis are unclear. Here we show that the microbiome modulates neuronal activity in specific brain regions of male mice to regulate canonical stress responses and social behaviours. Social deviation in germ-free and antibiotic-treated mice is associated with elevated levels of the stress hormone corticosterone, which is primarily produced by activation of the hypothalamus\u2013pituitary\u2013adrenal (HPA) axis. Adrenalectomy, antagonism of glucocorticoid receptors, or pharmacological inhibition of corticosterone synthesis effectively corrects social deficits following microbiome depletion. Genetic ablation of glucocorticoid receptors in specific brain regions or chemogenetic inactivation of neurons in the paraventricular nucleus of the hypothalamus that produce corticotrophin-releasing hormone (CRH) reverse social impairments in antibiotic-treated mice. Conversely, specific activation of CRH-expressing neurons in the paraventricular nucleus induces social deficits in mice with a normal microbiome. Via microbiome profiling and in vivo selection, we identify a bacterial species, Enterococcus faecalis, that promotes social activity and reduces corticosterone levels in mice following social stress. These studies suggest that specific gut bacteria can restrain the activation of the HPA axis, and show that the microbiome can affect social behaviours through discrete neuronal circuits that mediate stress responses in the brain.", "doi": "10.1038/s41586-021-03669-y", "pmcid": "PMC8346519", "issn": "0028-0836", "publisher": "Nature Publishing Group", "publication": "Nature", "publication_date": "2021-07-15", "series_number": "7867", "volume": "595", "issue": "7867", "pages": "409-414" }, { "id": "authors:74gj8-w4649", "collection": "authors", "collection_id": "74gj8-w4649", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20200813-144241318", "type": "article", "title": "Metabolic multistability and hysteresis in a model aerobe-anaerobe microbiome community", "author": [ { "family_name": "Khazaei", "given_name": "Tahmineh", "orcid": "0000-0002-4743-2383", "clpid": "Khazaei-Tahmineh" }, { "family_name": "Williams", "given_name": "Rory L.", "orcid": "0000-0003-2605-5790", "clpid": "Williams-Rory-L" }, { "family_name": "Bogatyrev", "given_name": "Said R.", "orcid": "0000-0003-0486-9451", "clpid": "Bogatyrev-Said-R" }, { "family_name": "Doyle", "given_name": "John C.", "orcid": "0000-0002-1828-2486", "clpid": "Doyle-J-C" }, { "family_name": "Henry", "given_name": "Christopher S.", "orcid": "0000-0001-8058-9123", "clpid": "Henry-Christopher-S" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Major changes in the microbiome are associated with health and disease. Some microbiome states persist despite seemingly unfavorable conditions, such as the proliferation of aerobe-anaerobe communities in oxygen-exposed environments in wound infections or small intestinal bacterial overgrowth. Mechanisms underlying transitions into and persistence of these states remain unclear. Using two microbial taxa relevant to the human microbiome, we combine genome-scale mathematical modeling, bioreactor experiments, transcriptomics, and dynamical systems theory to show that multistability and hysteresis (MSH) is a mechanism describing the shift from an aerobe-dominated state to a resilient, paradoxically persistent aerobe-anaerobe state. We examine the impact of changing oxygen and nutrient regimes and identify changes in metabolism and gene expression that lead to MSH and associated multi-stable states. In such systems, conceptual causation-correlation connections break and MSH must be used for analysis. Using MSH to analyze microbiome dynamics will improve our conceptual understanding of stability of microbiome states and transitions between states.", "doi": "10.1126/sciadv.aba0353", "pmcid": "PMC7423363", "issn": "2375-2548", "publisher": "American Association for the Advancement of Science", "publication": "Science Advances", "publication_date": "2020-08-12", "series_number": "33", "volume": "6", "issue": "33", "pages": "Art. No. eaba0353" }, { "id": "authors:6famx-f0654", "collection": "authors", "collection_id": "6famx-f0654", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20200303-103834329", "type": "article", "title": "A Quantitative Sequencing Framework for Absolute Abundance Measurements of Mucosal and Lumenal Microbial Communities", "author": [ { "family_name": "Barlow", "given_name": "Jacob T.", "orcid": "0000-0002-1842-4835", "clpid": "Barlow-Jacob-T" }, { "family_name": "Bogatyrev", "given_name": "Said R.", "orcid": "0000-0003-0486-9451", "clpid": "Bogatyrev-Said-R" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "A fundamental goal in microbiome studies is determining which microbes affect host physiology. Standard methods for determining changes in microbial taxa measure relative, rather than absolute abundances. Moreover, studies often analyze only stool, despite microbial diversity differing substantially among gastrointestinal (GI) locations. Here, we develop a quantitative framework to measure absolute abundances of individual bacterial taxa by combining the precision of digital PCR with the high-throughput nature of 16S rRNA gene amplicon sequencing. In a murine ketogenic-diet study, we compare microbial loads in lumenal and mucosal samples along the GI tract. Quantitative measurements of absolute (but not relative) abundances reveal decreases in total microbial loads on the ketogenic diet and enable us to determine the differential effects of diet on each taxon in stool and small-intestine mucosa samples. This rigorous quantitative microbial analysis framework, appropriate for diverse GI locations enables mapping microbial biogeography of the mammalian GI tract and more accurate analyses of changes in microbial taxa in microbiome studies.", "doi": "10.1038/s41467-020-16224-6", "pmcid": "PMC7244552", "issn": "2041-1723", "publisher": "Nature Publishing Group", "publication": "Nature Communications", "publication_date": "2020-05-22", "volume": "11", "pages": "Art. No. 2590" }, { "id": "authors:hh1ak-4sk21", "collection": "authors", "collection_id": "hh1ak-4sk21", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20200219-114918172", "type": "article", "title": "Self-partitioning SlipChip for slip-induced droplet formation and human papillomavirus viral load quantification with digital LAMP", "author": [ { "family_name": "Yu", "given_name": "Ziqing", "clpid": "Yu-Ziqing" }, { "family_name": "Lyu", "given_name": "Weiyuan", "clpid": "Lyu-Weiyuan" }, { "family_name": "Yu", "given_name": "Mengchao", "orcid": "0000-0002-2535-429X", "clpid": "Yu-Mengchao" }, { "family_name": "Wang", "given_name": "Qian", "clpid": "Wang-Qian" }, { "family_name": "Qu", "given_name": "Haijun", "clpid": "Qu-Haijun" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Han", "given_name": "Xu", "clpid": "Han-Xu" }, { "family_name": "Lai", "given_name": "Dongmei", "orcid": "0000-0003-3810-2136", "clpid": "Lai-Dongmei" }, { "family_name": "Shen", "given_name": "Feng", "orcid": "0000-0002-4709-330X", "clpid": "Shen-Feng" } ], "abstract": "Human papillomavirus (HPV) is one of the most common sexually transmitted infections worldwide, and persistent HPV infection can cause warts and even cancer. Nucleic acid analysis of HPV viral DNA can be very informative for the diagnosis and monitoring of HPV. Digital nucleic acid analysis, such as digital PCR and digital isothermal amplification, can provide sensitive detection and precise quantification of target nucleic acids, and its utility has been demonstrated in many biological research and medical diagnostic applications. A variety of methods have been developed for the generation of a large number of individual reaction partitions, a key requirement for digital nucleic acid analysis. However, an easily assembled and operated device for robust droplet formation without preprocessing devices, auxiliary instrumentation or control systems is still highly desired. In this paper, we present a self-partitioning SlipChip (sp-SlipChip) microfluidic device for the slip-induced generation of droplets to perform digital loop-mediated isothermal amplification (LAMP) for the detection and quantification of HPV DNA. In contrast to traditional SlipChip methods, which require the precise alignment of microfeatures, this sp-SlipChip utilized a design of \"chain-of-pearls\" continuous microfluidic channel that is independent of the overlapping of microfeatures on different plates to establish the fluidic path for reagent loading. Initiated by a simple slipping step, the aqueous solution can robustly self-partition into individual droplets by capillary pressure-driven flow. This advantage makes the sp-SlipChip very appealing for the point-of-care quantitative analysis of viral load. As a proof of concept, we performed digital LAMP on an sp-SlipChip to quantify human papillomaviruses (HPVs) 16 and 18 and tested this method with fifteen anonymous clinical samples.", "doi": "10.1016/j.bios.2020.112107", "issn": "0956-5663", "publisher": "Elsevier", "publication": "Biosensors and Bioelectronics", "publication_date": "2020-05-01", "volume": "155", "pages": "Art. No. 112107" }, { "id": "authors:e8d7z-m7780", "collection": "authors", "collection_id": "e8d7z-m7780", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20200302-155937790", "type": "article", "title": "Real-time kinetics and high-resolution melt curves in single-molecule digital LAMP to differentiate and study specific and non-specific amplification", "author": [ { "family_name": "Rolando", "given_name": "Justin C.", "orcid": "0000-0001-8948-319X", "clpid": "Rolando-Justin-C" }, { "family_name": "Jue", "given_name": "Erik", "orcid": "0000-0001-7585-3794", "clpid": "Jue-Erik" }, { "family_name": "Barlow", "given_name": "Jacob T.", "orcid": "0000-0002-1842-4835", "clpid": "Barlow-Jacob-T" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Isothermal amplification assays, such as loop-mediated isothermal amplification (LAMP), show great utility for the development of rapid diagnostics for infectious diseases because they have high sensitivity, pathogen-specificity and potential for implementation at the point of care. However, elimination of non-specific amplification remains a key challenge for the optimization of LAMP assays. Here, using chlamydia DNA as a clinically relevant target and high-throughput sequencing as an analytical tool, we investigate a potential mechanism of non-specific amplification. We then develop a real-time digital LAMP (dLAMP) with high-resolution melting temperature (HRM) analysis and use this single-molecule approach to analyze approximately 1.2 million amplification events. We show that single-molecule HRM provides insight into specific and non-specific amplification in LAMP that are difficult to deduce from bulk measurements. We use real-time dLAMP with HRM to evaluate differences between polymerase enzymes, the impact of assay parameters (e.g. time, rate or florescence intensity), and the effect background human DNA. By differentiating true and false positives, HRM enables determination of the optimal assay and analysis parameters that leads to the lowest limit of detection (LOD) in a digital isothermal amplification assay.", "doi": "10.1093/nar/gkaa099", "pmcid": "PMC7144905", "issn": "0305-1048", "publisher": "Oxford University Press", "publication": "Nucleic Acids Research", "publication_date": "2020-04-17", "series_number": "7", "volume": "48", "issue": "7", "pages": "Art. No. e42" }, { "id": "authors:zn887-tf656", "collection": "authors", "collection_id": "zn887-tf656", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20200320-065850359", "type": "article", "title": "Differential DNA accessibility to polymerase enables 30-minute phenotypic \u03b2-lactam antibiotic susceptibility testing of carbapenem-resistant Enterobacteriaceae", "author": [ { "family_name": "Schoepp", "given_name": "Nathan G.", "orcid": "0000-0002-2406-3693", "clpid": "Schoepp-Nathan-G" }, { "family_name": "Liaw", "given_name": "Eric J.", "orcid": "0000-0003-2244-8335", "clpid": "Liaw-Eric-J" }, { "family_name": "Winnett", "given_name": "Alexander", "orcid": "0000-0002-7338-5605", "clpid": "Winnett-A" }, { "family_name": "Savela", "given_name": "Emily S.", "orcid": "0000-0001-9614-4276", "clpid": "Savela-Emily-S" }, { "family_name": "Garner", "given_name": "Omai B.", "orcid": "0000-0002-7366-2692", "clpid": "Garner-Omai-B" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "The rise in carbapenem-resistant Enterobacteriaceae (CRE) infections has created a global health emergency, underlining the critical need to develop faster diagnostics to treat swiftly and correctly. Although rapid pathogen-identification (ID) tests are being developed, gold-standard antibiotic susceptibility testing (AST) remains unacceptably slow (1\u20132 d), and innovative approaches for rapid phenotypic ASTs for CREs are urgently needed. Motivated by this need, in this manuscript we tested the hypothesis that upon treatment with \u03b2-lactam antibiotics, susceptible Enterobacteriaceae isolates would become sufficiently permeabilized, making some of their DNA accessible to added polymerase and primers. Further, we hypothesized that this accessible DNA would be detectable directly by isothermal amplification methods that do not fully lyse bacterial cells. We build on these results to develop the polymerase-accessibility AST (pol-aAST), a new phenotypic approach for \u03b2-lactams, the major antibiotic class for gram-negative infections. We test isolates of the 3 causative pathogens of CRE infections using ceftriaxone (CRO), ertapenem (ETP), and meropenem (MEM) and demonstrate agreement with gold-standard AST. Importantly, pol-aAST correctly categorized resistant isolates that are undetectable by current genotypic methods (negative for \u03b2-lactamase genes or lacking predictive genotypes). We also test contrived and clinical urine samples. We show that the pol-aAST can be performed in 30 min sample-to-answer using contrived urine samples and has the potential to be performed directly on clinical urine specimens.", "doi": "10.1371/journal.pbio.3000652", "pmcid": "PMC7081982", "issn": "1545-7885", "publisher": "Public Library of Science", "publication": "PLoS Biology", "publication_date": "2020-03-19", "series_number": "3", "volume": "18", "issue": "3", "pages": "Art. No. 3000652" }, { "id": "authors:fhhh2-xg437", "collection": "authors", "collection_id": "fhhh2-xg437", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20200319-141031875", "type": "article", "title": "Surfactant-enhanced DNA accessibility to nuclease accelerates phenotypic \u03b2-lactam antibiotic susceptibility testing of Neisseria gonorrhoeae", "author": [ { "family_name": "Savela", "given_name": "Emily S.", "orcid": "0000-0001-9614-4276", "clpid": "Savela-E-S" }, { "family_name": "Schoepp", "given_name": "Nathan G.", "orcid": "0000-0002-2406-3693", "clpid": "Schoepp-N-G" }, { "family_name": "Cooper", "given_name": "Matthew M.", "orcid": "0000-0002-5868-5159", "clpid": "Cooper-M-M" }, { "family_name": "Rolando", "given_name": "Justin C.", "orcid": "0000-0001-8948-319X", "clpid": "Rolando-J-C" }, { "family_name": "Klausner", "given_name": "Jeffrey D.", "orcid": "0000-0002-6922-7364", "clpid": "Klausner-J-D" }, { "family_name": "Soge", "given_name": "Olusegun O.", "orcid": "0000-0002-8504-2319", "clpid": "Soge-O-O" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Rapid antibiotic susceptibility testing (AST) for Neisseria gonorrhoeae (Ng) is critically needed to counter widespread antibiotic resistance. Detection of nucleic acids in genotypic AST can be rapid, but it has not been successful for \u03b2-lactams (the largest antibiotic class used to treat Ng). Rapid phenotypic AST for Ng is challenged by the pathogen's slow doubling time and the lack of methods to quickly quantify the pathogen's response to \u03b2-lactams. Here, we asked two questions: (1) Is it possible to use nucleic acid quantification to measure the \u03b2-lactam susceptibility phenotype of Ng very rapidly, using antibiotic-exposure times much shorter than the 1- to 2-h doubling time of Ng? (2) Would such short-term antibiotic exposures predict the antibiotic resistance profile of Ng measured by plate growth assays over multiple days? To answer these questions, we devised an innovative approach for performing a rapid phenotypic AST that measures DNA accessibility to exogenous nucleases after exposure to \u03b2-lactams (termed nuclease-accessibility AST [nuc-aAST]). We showed that DNA in antibiotic-susceptible cells has increased accessibility upon exposure to \u03b2-lactams and that a judiciously chosen surfactant permeabilized the outer membrane and enhanced this effect. We tested penicillin, cefixime, and ceftriaxone and found good agreement between the results of the nuc-aAST after 15\u201330 min of antibiotic exposure and the results of the gold-standard culture-based AST measured over days. These results provide a new pathway toward developing a critically needed phenotypic AST for Ng and additional global-health threats.", "doi": "10.1371/journal.pbio.3000651", "pmcid": "PMC7081974", "issn": "1545-7885", "publisher": "Public Library of Science", "publication": "PLoS Biology", "publication_date": "2020-03-19", "series_number": "3", "volume": "18", "issue": "3", "pages": "Art. No. 3000651" }, { "id": "authors:ahsp6-e8a25", "collection": "authors", "collection_id": "ahsp6-e8a25", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20200212-152816065", "type": "article", "title": "Self-reinoculation with fecal flora changes microbiota density and composition leading to an altered bile-acid profile in the mouse small intestine", "author": [ { "family_name": "Bogatyrev", "given_name": "Said R.", "orcid": "0000-0003-0486-9451", "clpid": "Bogatyrev-S-R" }, { "family_name": "Rolando", "given_name": "Justin C.", "orcid": "0000-0001-8948-319X", "clpid": "Rolando-J-C" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Background: The upper gastrointestinal tract plays a prominent role in human physiology as the primary site for enzymatic digestion and nutrient absorption, immune sampling, and drug uptake. Alterations to the small intestine microbiome have been implicated in various human diseases, such as non-alcoholic steatohepatitis and inflammatory bowel conditions. Yet, the physiological and functional roles of the small intestine microbiota in humans remain poorly characterized because of the complexities associated with its sampling. Rodent models are used extensively in microbiome research and enable the spatial, temporal, compositional, and functional interrogation of the gastrointestinal microbiota and its effects on the host physiology and disease phenotype. Classical, culture-based studies have documented that fecal microbial self-reinoculation (via coprophagy) affects the composition and abundance of microbes in the murine proximal gastrointestinal tract. This pervasive self-reinoculation behavior could be a particularly relevant study factor when investigating small intestine microbiota. Modern microbiome studies either do not take self-reinoculation into account, or assume that approaches such as single housing mice or housing on wire mesh floors eliminate it. These assumptions have not been rigorously tested with modern tools. Here, we used quantitative 16S rRNA gene amplicon sequencing, quantitative microbial functional gene content inference, and metabolomic analyses of bile acids to evaluate the effects of self-reinoculation on microbial loads, composition, and function in the murine upper gastrointestinal tract. \n\nResults: In coprophagic mice, continuous self-exposure to the fecal flora had substantial quantitative and qualitative effects on the upper gastrointestinal microbiome. These differences in microbial abundance and community composition were associated with an altered profile of the small intestine bile acid pool, and, importantly, could not be inferred from analyzing large intestine or stool samples. Overall, the patterns observed in the small intestine of non-coprophagic mice (reduced total microbial load, low abundance of anaerobic microbiota, and bile acids predominantly in the conjugated form) resemble those typically seen in the human small intestine. \n\nConclusions: Future studies need to take self-reinoculation into account when using mouse models to evaluate gastrointestinal microbial colonization and function in relation to xenobiotic transformation and pharmacokinetics or in the context of physiological states and diseases linked to small intestine microbiome and to small intestine dysbiosis.", "doi": "10.1186/s40168-020-0785-4", "pmcid": "PMC7017497", "issn": "2049-2618", "publisher": "BioMed Central", "publication": "Microbiome", "publication_date": "2020-02-12", "volume": "8", "pages": "Art. No. 19" }, { "id": "authors:gjzvx-agj04", "collection": "authors", "collection_id": "gjzvx-agj04", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20200206-125057522", "type": "article", "title": "Two-phase wash to solve the ubiquitous contaminant-carryover problem in commercial nucleic-acid extraction kits", "author": [ { "family_name": "Jue", "given_name": "Erik", "orcid": "0000-0001-7585-3794", "clpid": "Jue-Erik" }, { "family_name": "Witters", "given_name": "Daan", "orcid": "0000-0003-2179-5300", "clpid": "Witter-Dann" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "The success of fundamental and applied nucleic acid (NA) research depends on NA purity, but obtaining pure NAs from raw, unprocessed samples is challenging. Purification using solid-phase NA extractions utilizes sequential additions of lysis and wash buffers followed by elution. The resulting eluent contains NAs and carryover of extraction buffers. Typically, these inhibitory buffers are heavily diluted by the reaction mix (e.g., 10x dilution is 1\u2009\u00b5L eluent in 9\u2009\u00b5L reaction mix), but in applications requiring high sensitivity (e.g., single-cell sequencing, pathogen diagnostics) it is desirable to use low dilutions (e.g., 2x) to maximize NA concentration. Here, we demonstrate pervasive carryover of inhibitory buffers into eluent when several commercial sample-preparation kits are used following manufacturer protocols. At low eluent dilution (2\u20132.5x) we observed significant reaction inhibition of polymerase chain reaction (PCR), loop-mediated isothermal amplification (LAMP), and reverse transcription (RT). We developed a two-phase wash (TPW) method by adding a wash buffer with low water solubility prior to the elution step. The TPW reduces carryover of extraction buffers, phase-separates from the eluent, and does not reduce NA yield (measured by digital PCR). We validated the TPW for silica columns and magnetic beads by demonstrating significant improvements in performance and reproducibility of qPCR, LAMP, and RT reactions.", "doi": "10.1038/s41598-020-58586-3", "pmcid": "PMC7004994", "issn": "2045-2322", "publisher": "Nature Publishing Group", "publication": "Scientific Reports", "publication_date": "2020-02-06", "volume": "10", "pages": "Art. No. 1940" }, { "id": "authors:91xgk-csy72", "collection": "authors", "collection_id": "91xgk-csy72", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20190823-090755927", "type": "article", "title": "Microfluidic SlipChip device for multistep multiplexed biochemistry on a nanoliter scale", "author": [ { "family_name": "Zhukov", "given_name": "Dmitriy V.", "orcid": "0000-0002-4834-3147", "clpid": "Zhukov-D-V" }, { "family_name": "Khorosheva", "given_name": "Eugenia M.", "orcid": "0000-0003-3620-4884", "clpid": "Khorosheva-E-M" }, { "family_name": "Khazaei", "given_name": "Tahmineh", "orcid": "0000-0002-4743-2383", "clpid": "Khazaei-Tahmineh" }, { "family_name": "Du", "given_name": "Wenbin", "orcid": "0000-0002-7401-1410", "clpid": "Du-Wenbin" }, { "family_name": "Selck", "given_name": "David A.", "clpid": "Selck-D-A" }, { "family_name": "Shishkin", "given_name": "Alexander A.", "clpid": "Shishkin-A-A" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "We have developed a multistep microfluidic device that expands the current SlipChip capabilities by enabling multiple steps of droplet merging and multiplexing. Harnessing the interfacial energy between carrier and sample phases, this manually operated device accurately meters nanoliter volumes of reagents and transfers them into on-device reaction wells. Judiciously shaped microfeatures and surface-energy traps merge droplets in a parallel fashion. Wells can be tuned for different volumetric capacities and reagent types, including for pre-spotted reagents that allow for unique identification of original well contents even after their contents are pooled. We demonstrate the functionality of the multistep SlipChip by performing RNA transcript barcoding on-device for synthetic spiked-in standards and for biologically derived samples. This technology is a good candidate for a wide range of biological applications that require multiplexing of multistep reactions in nanoliter volumes, including single-cell analyses.", "doi": "10.1039/c9lc00541b", "issn": "1473-0197", "publisher": "Royal Society of Chemistry", "publication": "Lab on a Chip", "publication_date": "2019-10-07", "series_number": "19", "volume": "19", "issue": "19", "pages": "3200-3211" }, { "id": "authors:j7xv9-9wj47", "collection": "authors", "collection_id": "j7xv9-9wj47", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20190823-085722494", "type": "article", "title": "Interplay of motility and polymer-driven depletion forces in the initial stages of bacterial aggregation", "author": [ { "family_name": "Porter", "given_name": "Michael K.", "orcid": "0000-0002-0777-7563", "clpid": "Porter-M-K" }, { "family_name": "Steinberg", "given_name": "Asher Preska", "orcid": "0000-0002-8694-7224", "clpid": "Steinberg-A-P" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Motile bacteria are often found in complex, polymer-rich environments in which microbes can aggregate via polymer-induced depletion forces. Bacterial aggregation has many biological implications; it can promote biofilm formation, upregulate virulence factors, and lead to quorum sensing. The steady state aggregation behavior of motile bacteria in polymer solutions has been well studied and shows that stronger depletion forces are required to aggregate motile bacteria as compared with their nonmotile analogs. However, no one has studied whether these same trends hold at the initial stages of aggregation. We use experiments and numerical calculations to investigate the polymer-induced depletion aggregation of motile Escherichia coli in polyethylene glycol solutions on short experimental timescales (\u223c10 min). Our work reveals that in the semi-dilute polymer concentration regime and at short timescales, in contrast to what is found at steady state, bacterial motility actually enhances aggregate formation by increasing the collision rate in viscous environments. These unexpected findings have implications for developing models of active matter, and for understanding bacterial aggregation in dynamic, biological environments, where the system may never reach steady state.", "doi": "10.1039/c9sm00791a", "issn": "1744-683X", "publisher": "Royal Society of Chemistry", "publication": "Soft Matter", "publication_date": "2019-09-21", "series_number": "35", "volume": "15", "issue": "35", "pages": "7071-7079" }, { "id": "authors:bwz2h-8h679", "collection": "authors", "collection_id": "bwz2h-8h679", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20190523-142111730", "type": "article", "title": "Multistep SlipChip for the Generation of Serial Dilution Nanoliter Arrays and Hepatitis B Viral Load Quantification by Digital Loop Mediated Isothermal Amplification", "author": [ { "family_name": "Yu", "given_name": "Mengchao", "orcid": "0000-0002-2535-429X", "clpid": "Yu-Mengchao" }, { "family_name": "Chen", "given_name": "Xiaoying", "clpid": "Chen-Xiaoying" }, { "family_name": "Qu", "given_name": "Haijun", "clpid": "Qu-Haijun" }, { "family_name": "Ma", "given_name": "Liang", "clpid": "Ma-Liang" }, { "family_name": "Xu", "given_name": "Lei", "clpid": "Xu-Lei" }, { "family_name": "Lv", "given_name": "Weiyuan", "clpid": "Lv-Weiyuan" }, { "family_name": "Wang", "given_name": "Hua", "clpid": "Wang-Hua" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Li", "given_name": "Min", "clpid": "Li-Min" }, { "family_name": "Shen", "given_name": "Feng", "orcid": "0000-0002-4709-330X", "clpid": "Shen-Feng" } ], "abstract": "Serial dilution is a commonly used technique that generates a low-concentration working sample from a high-concentration stock solution and is used to set up screening conditions over a large dynamic range for biological study, optimization of reaction conditions, drug screening, etc. Creating an array of serial dilutions usually requires cumbersome manual pipetting steps or a robotic liquid handling system. Moreover, it is very challenging to set up an array of serial dilutions in nanoliter volumes in miniaturized assays. Here, a multistep SlipChip microfluidic device is presented for generating serial dilution nanoliter arrays in high throughput with a series of simple sliding motions. The dilution ratio can be precisely predetermined by the volumes of mother microwells and daughter microwells, and this paper demonstrates devices designed to have dilution ratios of 1:1, 1:2, and 1:4. Furthermore, an eight-step serial dilution SlipChip with a dilution ratio of 1:4 is applied for digital loop-mediated isothermal amplification (LAMP) across a large dynamic range and tested for hepatitis B viral load quantification with clinical samples. With 64 wells of each dilution and fewer than 600 wells in total, the serial dilution SlipChip can achieve a theoretical quantification dynamic range of 7 orders of magnitude.", "doi": "10.1021/acs.analchem.9b01270", "issn": "0003-2700", "publisher": "American Chemical Society", "publication": "Analytical Chemistry", "publication_date": "2019-07-16", "series_number": "14", "volume": "91", "issue": "14", "pages": "8751-8755" }, { "id": "authors:0hjjc-kj831", "collection": "authors", "collection_id": "0hjjc-kj831", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20190619-091518942", "type": "article", "title": "Food Polyelectrolytes Compress the Colonic Mucus Hydrogel by a Donnan Mechanism", "author": [ { "family_name": "Preska Steinberg", "given_name": "Asher", "orcid": "0000-0002-8694-7224", "clpid": "Preska-Steinberg-A" }, { "family_name": "Wang", "given_name": "Zhen-Gang", "orcid": "0000-0002-3361-6114", "clpid": "Wang-Zhen-Gang" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Systems consisting of a polyelectrolyte solution in contact with a cross-linked polyelectrolyte network are ubiquitous (e.g., biofilms, drug-delivering hydrogels, and mammalian extracellular matrices), yet the underlying physics governing these interactions is not well understood. Here, we find that carboxymethyl cellulose, a polyelectrolyte commonly found in processed foods and associated with inflammation and obesity, compresses the colonic mucus hydrogel (a key regulator of host\u2013microbe interactions and a protective barrier) in mice. The extent of this polyelectrolyte-induced compression is enhanced by the degree of polymer negative charge. Through animal experiments and numerical calculations, we find that this phenomenon can be described by a Donnan mechanism. Further, the observed behavior can be quantitatively described by a simple, one-parameter model. This work suggests that polymer charge should be considered when developing food products because of its potential role in modulating the protective properties of colonic mucus.", "doi": "10.1021/acs.biomac.9b00442", "issn": "1525-7797", "publisher": "American Chemical Society", "publication": "Biomacromolecules", "publication_date": "2019-07-08", "series_number": "7", "volume": "20", "issue": "7", "pages": "2675-2683" }, { "id": "authors:k10ak-7tt92", "collection": "authors", "collection_id": "k10ak-7tt92", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20190710-125335706", "type": "article", "title": "Human-gut-microbiome on a chip", "author": [ { "family_name": "Poceviciute", "given_name": "Roberta", "clpid": "Poceviciute-R" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "A microfluidic chip incorporating oxygen gradients, a diverse human microbiota and patient-derived cells, mimics interactions between microorganisms and host tissue in the human gut.", "doi": "10.1038/s41551-019-0425-0", "issn": "2157-846X", "publisher": "Nature Publishing Group", "publication": "Nature Biomedical Engineering", "publication_date": "2019-07", "series_number": "7", "volume": "3", "issue": "7", "pages": "500-501" }, { "id": "authors:xq49y-sfb72", "collection": "authors", "collection_id": "xq49y-sfb72", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20200127-080528197", "type": "article", "title": "Cell Envelope Damage of N. gonorrhoeae after 15-min Beta-Lactam Exposure Enables Rapid Antimicrobial Susceptibility Testing", "author": [ { "family_name": "Savela", "given_name": "Emily", "clpid": "Savela-E" }, { "family_name": "Schoepp", "given_name": "Nathan G.", "orcid": "0000-0002-2406-3693", "clpid": "Schoepp-N-G" }, { "family_name": "Rolando", "given_name": "Justin", "orcid": "0000-0001-8948-319X", "clpid": "Rolando-J-C" }, { "family_name": "Soge", "given_name": "Olusegun", "clpid": "Soge-O" }, { "family_name": "Ismagilov", "given_name": "Rustem", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Background: Designing diagnostic tools to perform phenotypic antimicrobial susceptibility testing (AST) at the point-of-care (POC) is a vital step in tackling the global threat of antimicrobial resistance. Gonorrhea infections with resistance to the first-line dual therapy have already emerged, highlighting the impending threat of untreatable gonorrhea. A rapid, phenotypic AST could enable evidence-based (instead of empirical) therapy and improve surveillance. The focus of this work is to develop innovative strategies to measure the phenotypic antimicrobial susceptibility of Neisseria gonorrhoeae clinical isolates after just 15\u201330 min of exposure with an antibiotic. We focused on the duration of the exposure step because it remains the bottleneck for phenotypic AST with fastidious and slow-growing microorganisms. \n\nMethods: We selected nucleic acid readout because our longterm goals include building fully integrated POC devices that determine the phenotypic response to antibiotic of a specific pathogen rapidly. We have been developing rapid phenotypic ASTs based on quantification of nucleic-acid concentrations in antibiotic-exposed samples. We describe a new phenotypic AST that does not depend on the speed of DNA replication and applies to beta-lactams penicillin, ceftriaxone, and cefixime acting on clinical isolates of N. gonorrhoeae very rapidly. \n\nResults: Our assay had 100% categorical agreement with the gold-standard agar dilution AST when N. gonorrhoeae isolates were incubated for 15-min with penicillin, and 100% categorical agreement when incubated for 30 min with ceftriaxone and cefixime, and steps can be performed within 35 min measured from contrived urine samples exposed to penicillin. \n\nConclusion: By designing techniques which allow us to rapidly determine the antibiotic phenotype, evidence-based prescription of antibiotics will become possible.", "doi": "10.1136/sextrans-2019-sti.729", "issn": "1368-4973", "publisher": "BMJ Publishing Group", "publication": "Sexually Transmitted Infections", "publication_date": "2019-07", "series_number": "S1", "volume": "95", "issue": "S1", "pages": "A291" }, { "id": "authors:yvcat-rz898", "collection": "authors", "collection_id": "yvcat-rz898", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20181210-140412486", "type": "article", "title": "High-molecular-weight polymers from dietary fiber drive aggregation of particulates in the murine small intestine", "author": [ { "family_name": "Preska Steinberg", "given_name": "Asher", "orcid": "0000-0002-8694-7224", "clpid": "Preska-Steinberg-Asher" }, { "family_name": "Datta", "given_name": "Sujit S.", "orcid": "0000-0003-2400-1561", "clpid": "Datta-Sujit-S" }, { "family_name": "Naragon", "given_name": "Thomas", "orcid": "0000-0002-5373-4257", "clpid": "Naragon-Thomas-H" }, { "family_name": "Rolando", "given_name": "Justin C.", "orcid": "0000-0001-8948-319X", "clpid": "Rolando-Justin-C" }, { "family_name": "Bogatyrev", "given_name": "Said R.", "orcid": "0000-0003-0486-9451", "clpid": "Bogatyrev-Said-R" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "The lumen of the small intestine (SI) is filled with particulates: microbes, therapeutic particles, and food granules. The structure of this particulate suspension could impact uptake of drugs and nutrients and the function of microorganisms; however, little is understood about how this suspension is re-structured as it transits the gut. Here, we demonstrate that particles spontaneously aggregate in SI luminal fluid ex vivo. We find that mucins and immunoglobulins are not required for aggregation. Instead, aggregation can be controlled using polymers from dietary fiber in a manner that is qualitatively consistent with polymer-induced depletion interactions, which do not require specific chemical interactions. Furthermore, we find that aggregation is tunable; by feeding mice dietary fibers of different molecular weights, we can control aggregation in SI luminal fluid. This work suggests that the molecular weight and concentration of dietary polymers play an underappreciated role in shaping the physicochemical environment of the gut.", "doi": "10.7554/eLife.40387", "pmcid": "PMC6342521", "issn": "2050-084X", "publisher": "eLife Sciences Publications", "publication": "eLife", "publication_date": "2019-01-22", "volume": "8", "pages": "Art. No. e40387" }, { "id": "authors:dk2yf-f8095", "collection": "authors", "collection_id": "dk2yf-f8095", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20181221-105356677", "type": "article", "title": "Real-Time, Digital LAMP with Commercial Microfluidic Chips Reveals the Interplay of Efficiency, Speed, and Background Amplification as a Function of Reaction Temperature and Time", "author": [ { "family_name": "Rolando", "given_name": "Justin C.", "orcid": "0000-0001-8948-319X", "clpid": "Rolando-Justin-C" }, { "family_name": "Jue", "given_name": "Erik", "orcid": "0000-0001-7585-3794", "clpid": "Jue-Erik" }, { "family_name": "Schoepp", "given_name": "Nathan G.", "orcid": "0000-0002-2406-3693", "clpid": "Schoepp-Nathan-G" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Real-time, isothermal, digital nucleic acid amplification is emerging as an attractive approach for a multitude of applications including diagnostics, mechanistic studies, and assay optimization. Unfortunately, there is no commercially available and affordable real-time, digital instrument validated for isothermal amplification; thus, most researchers have not been able to apply digital, real-time approaches to isothermal amplification. Here, we generate an approach to real-time digital loop-mediated isothermal amplification (LAMP) using commercially available microfluidic chips and reagents and open-source components. We demonstrate this approach by testing variables that influence LAMP reaction speed and the probability of detection. By analyzing the interplay of amplification efficiency, background, and speed of amplification, this real-time digital method enabled us to test enzymatic performance over a range of temperatures, generating high-precision kinetic and end-point measurements. We were able to identify the unique optimal temperature for two polymerase enzymes while accounting for amplification efficiency, nonspecific background, and time to threshold. We validated this digital LAMP assay and pipeline by performing a phenotypic antibiotic susceptibility test on 17 archived clinical urine samples from patients diagnosed with urinary tract infections. We provide all the necessary workflows to perform digital LAMP using standard laboratory equipment and commercially available materials. This real-time digital approach will be useful to others in the future to understand the fundamentals of isothermal chemistries, including which components determine amplification fate, reaction speed, and enzymatic performance. Researchers can also adapt this pipeline, which uses only standard equipment and commercial components, to quickly study and optimize assays using precise, real-time digital quantification, accelerating development of critically needed diagnostics.", "doi": "10.1021/acs.analchem.8b04324", "pmcid": "PMC6322147", "issn": "0003-2700", "publisher": "American Chemical Society", "publication": "Analytical Chemistry", "publication_date": "2019-01-02", "series_number": "1", "volume": "91", "issue": "1", "pages": "1034-1042" }, { "id": "authors:jhqfe-dfs86", "collection": "authors", "collection_id": "jhqfe-dfs86", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20180802-093834589", "type": "article", "title": "RNA markers enable phenotypic test of antibiotic susceptibility in Neisseria gonorrhoeae after 10\u2009minutes of ciprofloxacin exposure", "author": [ { "family_name": "Khazaei", "given_name": "Tahmineh", "orcid": "0000-0002-4743-2383", "clpid": "Khazaei-Tahmineh" }, { "family_name": "Barlow", "given_name": "Jacob T.", "orcid": "0000-0002-1842-4835", "clpid": "Barlow-J-T" }, { "family_name": "Schoepp", "given_name": "Nathan G.", "orcid": "0000-0002-2406-3693", "clpid": "Schoepp-N-G" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Antimicrobial-resistant Neisseria gonorrhoeae is an urgent public-health threat, with continued worldwide incidents of infection and rising resistance to antimicrobials. Traditional culture-based methods for antibiotic susceptibility testing are unacceptably slow (1\u20132 days), resulting in the use of broad-spectrum antibiotics and the further development and spread of resistance. Critically needed is a rapid antibiotic susceptibility test (AST) that can guide treatment at the point-of-care. Rapid phenotypic approaches using quantification of DNA have been demonstrated for fast-growing organisms (e.g. E. coli) but are challenging for slower-growing pathogens such as N. gonorrhoeae. Here, we investigate the potential of RNA signatures to provide phenotypic responses to antibiotics in N. gonorrhoeae that are faster and greater in magnitude compared with DNA. Using RNA sequencing, we identified antibiotic-responsive transcripts. Significant shifts (>4-fold change) in transcript levels occurred within 5\u2009min of antibiotic exposure. We designed assays for responsive transcripts with the highest abundances and fold changes, and validated gene expression using digital PCR. Using the top two markers (porB and rpmB) we correctly determined the antibiotic susceptibility and resistance of 49 clinical isolates after 10\u2009min exposure to ciprofloxacin. RNA signatures are therefore promising as an approach on which to build rapid AST devices for N. gonorrhoeae at the point-of-care, which is critical for disease management, surveillance, and antibiotic stewardship efforts.", "doi": "10.1038/s41598-018-29707-w", "pmcid": "PMC6072703", "issn": "2045-2322", "publisher": "Nature Publishing Group", "publication": "Scientific Reports", "publication_date": "2018-08-02", "volume": "8", "pages": "Art. No. 11606" }, { "id": "authors:ye1wh-40d17", "collection": "authors", "collection_id": "ye1wh-40d17", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20161213-131157930", "type": "article", "title": "Rapid pathogen-specific phenotypic antibiotic susceptibility testing using digital LAMP quantification in clinical samples", "author": [ { "family_name": "Schoepp", "given_name": "Nathan G.", "orcid": "0000-0002-2406-3693", "clpid": "Schoepp-Nathan-G" }, { "family_name": "Schlappi", "given_name": "Travis S.", "orcid": "0000-0001-6132-6459", "clpid": "Schlappi-Travis-S" }, { "family_name": "Curtis", "given_name": "Matthew S.", "orcid": "0000-0002-9662-3266", "clpid": "Curtis-Matthew-S" }, { "family_name": "Butkovich", "given_name": "Slava S.", "orcid": "0000-0001-9468-595X", "clpid": "Butkovich-Slava-S" }, { "family_name": "Miller", "given_name": "Shelley", "clpid": "Miller-Shelley" }, { "family_name": "Humphries", "given_name": "Romney M.", "orcid": "0000-0002-6568-156X", "clpid": "Humphries-Romney-M" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Rapid antimicrobial susceptibility testing (AST) is urgently needed for informing treatment decisions and preventing the spread of antimicrobial resistance resulting from the misuse and overuse of antibiotics. To date, no phenotypic AST exists that can be performed within a single patient visit (30 min) directly from clinical samples. We show that AST results can be obtained by using digital nucleic acid quantification to measure the phenotypic response of Escherichia coli present within clinical urine samples exposed to an antibiotic for 15 min. We performed this rapid AST using our ultrafast (~7 min) digital real-time loop-mediated isothermal amplification (dLAMP) assay [area under the curve (AUC), 0.96] and compared the results to a commercial (~2 hours) digital polymerase chain reaction assay (AUC, 0.98). The rapid dLAMP assay can be used with SlipChip microfluidic devices to determine the phenotypic antibiotic susceptibility of E. coli directly from clinical urine samples in less than 30 min. With further development for additional pathogens, antibiotics, and sample types, rapid digital AST (dAST) could enable rapid clinical decision-making, improve management of infectious diseases, and facilitate antimicrobial stewardship.", "doi": "10.1126/scitranslmed.aal3693", "pmcid": "PMC6765391", "issn": "1946-6234", "publisher": "American Association for the Advancement of Science", "publication": "Science Translational Medicine", "publication_date": "2017-10-04", "series_number": "410", "volume": "9", "issue": "410", "pages": "Art. No. eaal3693" }, { "id": "authors:kmb2q-cjq59", "collection": "authors", "collection_id": "kmb2q-cjq59", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20170628-083437438", "type": "article", "title": "Conceptual and Experimental Tools to Understand Spatial Effects and Transport Phenomena in Nonlinear Biochemical Networks Illustrated with Patchy Switching", "author": [ { "family_name": "Pompano", "given_name": "Rebecca R.", "orcid": "0000-0002-8644-9313", "clpid": "Pompano-Rebecca-R" }, { "family_name": "Chiang", "given_name": "Andrew H.", "clpid": "Chiang-Andrew-H" }, { "family_name": "Kastrup", "given_name": "Christian J.", "orcid": "0000-0003-3644-0019", "clpid": "Kastrup-Christian-J" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Many biochemical systems are spatially heterogeneous and exhibit nonlinear behaviors, such as state switching in response to small changes in the local concentration of diffusible molecules. Systems as varied as blood clotting, intracellular calcium signaling, and tissue inflammation are all heavily influenced by the balance of rates of reaction and mass transport phenomena including flow and diffusion. Transport of signaling molecules is also affected by geometry and chemoselective confinement via matrix binding. In this review, we use a phenomenon referred to as patchy switching to illustrate the interplay of nonlinearities, transport phenomena, and spatial effects. Patchy switching describes a change in the state of a network when the local concentration of a diffusible molecule surpasses a critical threshold. Using patchy switching as an example, we describe conceptual tools from nonlinear dynamics and chemical engineering that make testable predictions and provide a unifying description of the myriad possible experimental observations. We describe experimental microfluidic and biochemical tools emerging to test conceptual predictions by controlling transport phenomena and spatial distribution of diffusible signals, and we highlight the unmet need for in vivo tools.", "doi": "10.1146/annurev-biochem-060815-014207", "pmcid": "PMC10852032", "issn": "0066-4154", "publisher": "Annual Reviews", "publication": "Annual Review of Biochemistry", "publication_date": "2017-06", "volume": "86", "pages": "333-356" }, { "id": "authors:g9ews-vzq36", "collection": "authors", "collection_id": "g9ews-vzq36", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20161031-113924216", "type": "article", "title": "Instrument for Real-Time Digital Nucleic Acid Amplification on Custom Microfluidic Devices", "author": [ { "family_name": "Selck", "given_name": "David A.", "clpid": "Selck-D-A" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Nucleic acid amplification tests that are coupled with a digital readout enable the absolute quantification of single molecules, even at ultralow concentrations. Digital methods are robust, versatile and compatible with many amplification chemistries including isothermal amplification, making them particularly invaluable to assays that require sensitive detection, such as the quantification of viral load in occult infections or detection of sparse amounts of DNA from forensic samples. A number of microfluidic platforms are being developed for carrying out digital amplification. However, the mechanistic investigation and optimization of digital assays has been limited by the lack of real-time kinetic information about which factors affect the digital efficiency and analytical sensitivity of a reaction. Commercially available instruments that are capable of tracking digital reactions in real-time are restricted to only a small number of device types and sample-preparation strategies. Thus, most researchers who wish to develop, study, or optimize digital assays rely on the rate of the amplification reaction when performed in a bulk experiment, which is now recognized as an unreliable predictor of digital efficiency. To expand our ability to study how digital reactions proceed in real-time and enable us to optimize both the digital efficiency and analytical sensitivity of digital assays, we built a custom large-format digital real-time amplification instrument that can accommodate a wide variety of devices, amplification chemistries and sample-handling conditions. Herein, we validate this instrument, we provide detailed schematics that will enable others to build their own custom instruments, and we include a complete custom software suite to collect and analyze the data retrieved from the instrument. We believe assay optimizations enabled by this instrument will improve the current limits of nucleic acid detection and quantification, improving our fundamental understanding of single-molecule reactions and providing advancements in practical applications such as medical diagnostics, forensics and environmental sampling.", "doi": "10.1371/journal.pone.0163060", "pmcid": "PMC5070811", "issn": "1932-6203", "publisher": "Public Library of Science", "publication": "PLOS ONE", "publication_date": "2016-10", "series_number": "10", "volume": "11", "issue": "10", "pages": "Art. No. e0163060" }, { "id": "authors:kzer7-dy654", "collection": "authors", "collection_id": "kzer7-dy654", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20160705-083001025", "type": "article", "title": "Digital Quantification of DNA Replication and Chromosome Segregation Enables Determination of Antimicrobial Susceptibility after only 15 Minutes of Antibiotic Exposure", "author": [ { "family_name": "Schoepp", "given_name": "Nathan G.", "orcid": "0000-0002-2406-3693", "clpid": "Schoepp-Nathan-G" }, { "family_name": "Khorosheva", "given_name": "Eugenia M.", "orcid": "0000-0003-3620-4884", "clpid": "Khorosheva-Eugenia-M" }, { "family_name": "Schlappi", "given_name": "Travis S.", "orcid": "0000-0001-6132-6459", "clpid": "Schlappi-Travis-S" }, { "family_name": "Curtis", "given_name": "Matthew S.", "orcid": "0000-0002-9662-3266", "clpid": "Curtis-Matthew-S" }, { "family_name": "Humphries", "given_name": "Romney M.", "orcid": "0000-0002-6568-156X", "clpid": "Humphries-Romney-M" }, { "family_name": "Hindler", "given_name": "Janet A.", "clpid": "Hindler-Janet-A" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Rapid antimicrobial susceptibility testing (AST) would decrease misuse and overuse of antibiotics. The \"holy grail\" of AST is a phenotype-based test that can be performed within a doctor visit. Such a test requires the ability to determine a pathogen's susceptibility after only a short antibiotic exposure. Herein, digital PCR (dPCR) was employed to test whether measuring DNA replication of the target pathogen through digital single-molecule counting would shorten the required time of antibiotic exposure. Partitioning bacterial chromosomal DNA into many small volumes during dPCR enabled AST results after short exposure times by 1)\u2005precise quantification and 2)\u2005a measurement of how antibiotics affect the states of macromolecular assembly of bacterial chromosomes. This digital AST (dAST) determined susceptibility of clinical isolates from urinary tract infections (UTIs) after 15\u2005min of exposure for all four antibiotic classes relevant to UTIs. This work lays the foundation to develop a rapid, point-of-care AST and strengthen global antibiotic stewardship.", "doi": "10.1002/ange.201602763", "pmcid": "PMC5215780", "issn": "0570-0833", "publisher": "Wiley", "publication": "Angewandte Chemie International Edition in English", "publication_date": "2016-08-08", "series_number": "33", "volume": "55", "issue": "33", "pages": "9557-9561" }, { "id": "authors:2xw3m-2dy26", "collection": "authors", "collection_id": "2xw3m-2dy26", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20160725-102649276", "type": "article", "title": "Flow-through Capture and in Situ Amplification Can Enable Rapid Detection of a Few Single Molecules of Nucleic Acids from Several Milliliters of Solution", "author": [ { "family_name": "Schlappi", "given_name": "Travis S.", "orcid": "0000-0001-6132-6459", "clpid": "Schlappi-T-S" }, { "family_name": "McCalla", "given_name": "Stephanie E.", "clpid": "McCalla-S-E" }, { "family_name": "Schoepp", "given_name": "Nathan G.", "orcid": "0000-0002-2406-3693", "clpid": "Schoepp-N-G" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Detecting nucleic acids (NAs) at zeptomolar concentrations (few molecules per milliliter) currently requires expensive equipment and lengthy processing times to isolate and concentrate the NAs into a volume that is amenable to amplification processes, such as PCR or LAMP. Shortening the time required to concentrate NAs and integrating this procedure with amplification on-device would be invaluable to a number of analytical fields, including environmental monitoring and clinical diagnostics. Microfluidic point-of-care (POC) devices have been designed to address these needs, but they are not able to detect NAs present in zeptomolar concentrations in short time frames because they require slow flow rates and/or they are unable to handle milliliter-scale volumes. In this paper, we theoretically and experimentally investigate a flow-through capture membrane that solves this problem by capturing NAs with high sensitivity in a short time period, followed by direct detection via amplification. Theoretical predictions guided the choice of physical parameters for a chitosan-coated nylon membrane; these predictions can also be applied generally to other capture situations with different requirements. The membrane is also compatible with in situ amplification, which, by eliminating an elution step enables high sensitivity and will facilitate integration of this method into sample-to-answer detection devices. We tested a wide range of combinations of sample volumes and concentrations of DNA molecules using a capture membrane with a 2 mm radius. We show that for nucleic acid detection, this approach can concentrate and detect as few as \u223c10 molecules of DNA with flow rates as high as 1 mL/min, handling samples as large as 50 mL. In a specific example, this method reliably concentrated and detected \u223c25 molecules of DNA from 50 mL of sample.", "doi": "10.1021/acs.analchem.6b01485", "issn": "0003-2700", "publisher": "American Chemical Society", "publication": "Analytical Chemistry", "publication_date": "2016-08-02", "series_number": "15", "volume": "88", "issue": "15", "pages": "7647-7653" }, { "id": "authors:x2xza-km768", "collection": "authors", "collection_id": "x2xza-km768", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20160614-141541328", "type": "article", "title": "Polymers in the gut compress the colonic mucus hydrogel", "author": [ { "family_name": "Datta", "given_name": "Sujit S.", "orcid": "0000-0003-2400-1561", "clpid": "Datta-Sujit-S" }, { "family_name": "Preska Steinberg", "given_name": "Asher", "orcid": "0000-0002-8694-7224", "clpid": "Preska-Steinberg-Asher" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Colonic mucus is a key biological hydrogel that protects the gut from infection and physical damage and mediates host\u2013microbe interactions and drug delivery. However, little is known about how its structure is influenced by materials it comes into contact with regularly. For example, the gut abounds in polymers such as dietary fibers or administered therapeutics, yet whether such polymers interact with the mucus hydrogel, and if so, how, remains unclear. Although several biological processes have been identified as potential regulators of mucus structure, the polymeric composition of the gut environment has been ignored. Here, we demonstrate that gut polymers do in fact regulate mucus hydrogel structure, and that polymer\u2013mucus interactions can be described using a thermodynamic model based on Flory\u2013Huggins solution theory. We found that both dietary and therapeutic polymers dramatically compressed murine colonic mucus ex vivo and in vivo. This behavior depended strongly on both polymer concentration and molecular weight, in agreement with the predictions of our thermodynamic model. Moreover, exposure to polymer-rich luminal fluid from germ-free mice strongly compressed the mucus hydrogel, whereas exposure to luminal fluid from specific-pathogen-free mice\u2014whose microbiota degrade gut polymers\u2014did not; this suggests that gut microbes modulate mucus structure by degrading polymers. These findings highlight the role of mucus as a responsive biomaterial, and reveal a mechanism of mucus restructuring that must be integrated into the design and interpretation of studies involving therapeutic polymers, dietary fibers, and fiber-degrading gut microbes.", "doi": "10.1073/pnas.1602789113", "pmcid": "PMC4932961", "issn": "0027-8424", "publisher": "National Academy of Sciences", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "publication_date": "2016-06-28", "series_number": "26", "volume": "113", "issue": "26", "pages": "7041-7046" }, { "id": "authors:7jrba-0e405", "collection": "authors", "collection_id": "7jrba-0e405", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20160502-101837713", "type": "article", "title": "Evaluating 3D printing to solve the sample-to-device interface for LRS and POC diagnostics: example of an interlock meter-mix device for metering and lysing clinical urine samples", "author": [ { "family_name": "Jue", "given_name": "Erik", "orcid": "0000-0001-7585-3794", "clpid": "Jue-Erik" }, { "family_name": "Schoepp", "given_name": "Nathan G.", "orcid": "0000-0002-2406-3693", "clpid": "Schoepp-N-G" }, { "family_name": "Witters", "given_name": "Daan", "clpid": "Witters-D" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper evaluates the potential of 3D printing, a semi-automated additive prototyping technology, as a means to design and prototype a sample-to-device interface, amenable to diagnostics in limited-resource settings, where speed, accuracy and user-friendly design are critical components. As a test case, we built and validated an interlock meter-mix device for accurately metering and lysing human urine samples for use in downstream nucleic acid amplification. Two plungers and a multivalve generated and controlled fluid flow through the device and demonstrate the utility of 3D printing to create leak-free seals. Device operation consists of three simple steps that must be performed sequentially, eliminating manual pipetting and vortexing to provide rapid (5 to 10 s) and accurate metering and mixing. Bretherton's prediction was applied, using the bond number to guide a design that prevents potentially biohazardous samples from leaking from the device. We employed multi-material 3D printing technology, which allows composites with rigid and elastomeric properties to be printed as a single part. To validate the meter-mix device with a clinically relevant sample, we used urine spiked with inactivated Chlamydia trachomatis and Neisseria gonorrhoeae. A downstream nucleic acid amplification by quantitative PCR (qPCR) confirmed there was no statistically significant difference between samples metered and mixed using the standard protocol and those prepared with the meter-mix device, showing the 3D-printed device could accurately meter, mix and dispense a human urine sample without loss of nucleic acids. Although there are some limitations to 3D printing capabilities (e.g. dimension limitations related to support material used in the printing process), the advantages of customizability, modularity and rapid prototyping illustrate the utility of 3D printing for developing sample-to-device interfaces for diagnostics.", "doi": "10.1039/C6LC00292G", "issn": "1473-0197", "publisher": "Royal Society of Chemistry", "publication": "Lab on a Chip", "publication_date": "2016-05-21", "series_number": "10", "volume": "16", "issue": "10", "pages": "1852-1860" }, { "id": "authors:taq1z-0hn65", "collection": "authors", "collection_id": "taq1z-0hn65", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20160216-130203405", "type": "article", "title": "High throughput Single-cell Cultivation on Microfluidic Streak Plates", "author": [ { "family_name": "Jiang", "given_name": "Cheng-Ying", "clpid": "Jiang-Cheng-Ying" }, { "family_name": "Dong", "given_name": "Libing", "clpid": "Dong-Libing" }, { "family_name": "Zhao", "given_name": "Jian-Kang", "clpid": "Zhao-Jian-Kang" }, { "family_name": "Hu", "given_name": "Xiaofang", "clpid": "Hu-Xiaofang" }, { "family_name": "Shen", "given_name": "Chaohua", "clpid": "Shen-Chaohua" }, { "family_name": "Qiao", "given_name": "Yuxin", "clpid": "Qiao-Yuxin" }, { "family_name": "Zhang", "given_name": "Xinyue", "clpid": "Zhang-Xinyue" }, { "family_name": "Wang", "given_name": "Yapei", "clpid": "Wang-Yapei" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Liu", "given_name": "Shuang-Jiang", "clpid": "Liu-Shuang-Jiang" }, { "family_name": "Du", "given_name": "Wenbin", "orcid": "0000-0002-7401-1410", "clpid": "Du-Wenbin" } ], "abstract": "This paper describes the microfluidic streak plate (MSP), a facile method for high-throughput microbial cell separation and cultivation in nanoliter sessile droplets. The MSP method builds upon the conventional streak plate technique by using microfluidic devices to generate nanoliter droplets that can be streaked manually or robotically onto petri dishes prefilled with carrier oil for cultivation of single cells. In addition, chemical gradients could be encoded in the droplet array for comprehensive dose-response analysis. The MSP method was validated by using single-cell isolation of Escherichia coli and antimicrobial susceptibility testing of Pseudomonas aeruginosa PAO1. The robustness of the MSP work flow was demonstrated by cultivating a soil community that degrades polycyclic aromatic hydrocarbons. Cultivation in droplets enabled detection of the richest species diversity with better coverage of rare species. Moreover, isolation and cultivation of bacterial strains by MSP led to the discovery of several species with high degradation efficiency, including four Mycobacterium isolates and a previously unknown fluoranthene-degrading Blastococcus species.", "doi": "10.1128/AEM.03588-15", "pmcid": "PMC4807504", "issn": "0099-2240", "publisher": "American Society for Microbiology", "publication": "Applied and Environmental Microbiology", "publication_date": "2016-04", "series_number": "7", "volume": "82", "issue": "7", "pages": "2210-2218" }, { "id": "authors:fycvn-0ez10", "collection": "authors", "collection_id": "fycvn-0ez10", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20160223-132834891", "type": "article", "title": "Reading Out Single-Molecule Digital RNA and DNA Isothermal Amplification in Nanoliter Volumes with Unmodified Camera Phones", "author": [ { "family_name": "Rodriguez-Manzano", "given_name": "Jesus", "orcid": "0000-0002-2583-8366", "clpid": "Rodriguez-Manzano-Jesus" }, { "family_name": "Karymov", "given_name": "Mikhail A.", "clpid": "Karymov-Mikhail-A" }, { "family_name": "Begolo", "given_name": "Stefano", "clpid": "Begolo-Stefano" }, { "family_name": "Selck", "given_name": "David A.", "orcid": "0000-0002-0591-4165", "clpid": "Selck-David-A" }, { "family_name": "Zhukov", "given_name": "Dmitriy V.", "orcid": "0000-0002-4834-3147", "clpid": "Zhukov-Dmitriy-V" }, { "family_name": "Jue", "given_name": "Erik", "orcid": "0000-0001-7585-3794", "clpid": "Jue-Erik" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Digital single-molecule technologies are expanding diagnostic capabilities, enabling the ultrasensitive quantification of targets, such as viral load in HIV and hepatitis C infections, by directly counting single molecules. Replacing fluorescent readout with a robust visual readout that can be captured by any unmodified cell phone camera will facilitate the global distribution of diagnostic tests, including in limited-resource settings where the need is greatest. This paper describes a methodology for developing a visual readout system for digital single-molecule amplification of RNA and DNA by (i) selecting colorimetric amplification-indicator dyes that are compatible with the spectral sensitivity of standard mobile phones, and (ii) identifying an optimal ratiometric image-process for a selected dye to achieve a readout that is robust to lighting conditions and camera hardware and provides unambiguous quantitative results, even for colorblind users. We also include an analysis of the limitations of this methodology, and provide a microfluidic approach that can be applied to expand dynamic range and improve reaction performance, allowing ultrasensitive, quantitative measurements at volumes as low as 5 nL. We validate this methodology using SlipChip-based digital single-molecule isothermal amplification with \u03bbDNA as a model and hepatitis C viral RNA as a clinically relevant target. The innovative combination of isothermal amplification chemistry in the presence of a judiciously chosen indicator dye and ratiometric image processing with SlipChip technology allowed the sequence-specific visual readout of single nucleic acid molecules in nanoliter volumes with an unmodified cell phone camera. When paired with devices that integrate sample preparation and nucleic acid amplification, this hardware-agnostic approach will increase the affordability and the distribution of quantitative diagnostic and environmental tests.", "doi": "10.1021/acsnano.5b07338", "pmcid": "PMC4819493", "issn": "1936-0851", "publisher": "American Chemical Society", "publication": "ACS Nano", "publication_date": "2016-03", "series_number": "3", "volume": "10", "issue": "3", "pages": "3102-3113" }, { "id": "authors:563jp-mhk84", "collection": "authors", "collection_id": "563jp-mhk84", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20150918-141812518", "type": "article", "title": "Lack of correlation between reaction speed and analytical sensitivity in isothermal amplification reveals the value of digital methods for optimization: validation using digital real-time RT-LAMP", "author": [ { "family_name": "Khorosheva", "given_name": "Eugenia M.", "orcid": "0000-0003-3620-4884", "clpid": "Khorosheva-Eugenia-M" }, { "family_name": "Karymov", "given_name": "Mikhail A.", "clpid": "Karymov-Mikhail-A" }, { "family_name": "Selck", "given_name": "David A.", "orcid": "0000-0002-0591-4165", "clpid": "Selck-David-A" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "In this paper, we asked if it is possible to identify the best primers and reaction conditions based on improvements in reaction speed when optimizing isothermal reactions. We used digital single-molecule, real-time analyses of both speed and efficiency of isothermal amplification reactions, which revealed that improvements in the speed of isothermal amplification reactions did not always correlate with improvements in digital efficiency (the fraction of molecules that amplify) or with analytical sensitivity. However, we observed that the speeds of amplification for single-molecule (in a digital device) and multi-molecule (e.g. in a PCR well plate) formats always correlated for the same conditions. Also, digital efficiency correlated with the analytical sensitivity of the same reaction performed in a multi-molecule format. Our finding was supported experimentally with examples of primer design, the use or exclusion of loop primers in different combinations, and the use of different enzyme mixtures in one-step reverse-transcription loop-mediated amplification (RT-LAMP). Our results show that measuring the digital efficiency of amplification of single-template molecules allows quick, reliable comparisons of the analytical sensitivity of reactions under any two tested conditions, independent of the speeds of the isothermal amplification reactions.", "doi": "10.1093/nar/gkv877", "pmcid": "PMC4737171", "issn": "0305-1048", "publisher": "Oxford University Press", "publication": "Nucleic Acids Research", "publication_date": "2016-01-29", "series_number": "2", "volume": "44", "issue": "2", "pages": "Art. No. E10" }, { "id": "authors:ycm4f-g2c72", "collection": "authors", "collection_id": "ycm4f-g2c72", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20150623-114233631", "type": "article", "title": "A biochemical network can control formation of a synthetic material by sensing numerous specific stimuli", "author": [ { "family_name": "Yeon", "given_name": "Ju Hun", "clpid": "Yeon-Ju-Hun" }, { "family_name": "Chan", "given_name": "Karen Y. T.", "clpid": "Chan-Karen-Y-T" }, { "family_name": "Wong", "given_name": "Ting-Chia", "clpid": "Wong-Ting-Chia" }, { "family_name": "Chan", "given_name": "Kelvin", "clpid": "Chan-Kelvin" }, { "family_name": "Sutherland", "given_name": "Michael R.", "clpid": "Sutherland-M-R" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Pryzdial", "given_name": "Edward L. G.", "clpid": "Pryzdial-E-L-G" }, { "family_name": "Kastrup", "given_name": "Christian J.", "clpid": "Kastrup-C-J" } ], "abstract": "Developing bio-compatible smart materials that assemble in response to environmental cues requires strategies that can discriminate multiple specific stimuli in a complex milieu. Synthetic materials have yet to achieve this level of sensitivity, which would emulate the highly evolved and tailored reaction networks of complex biological systems. Here we show that the output of a naturally occurring network can be replaced with a synthetic material. Exploiting the blood coagulation system as an exquisite biological sensor, the fibrin clot end-product was replaced with a synthetic material under the biological control of a precisely regulated cross-linking enzyme. The functions of the coagulation network remained intact when the material was incorporated. Clot-like polymerization was induced in indirect response to distinct small molecules, phospholipids, enzymes, cells, viruses, an inorganic solid, a polyphenol, a polysaccharide, and a membrane protein. This strategy demonstrates for the first time that an existing stimulus-responsive biological network can be used to control the formation of a synthetic material by diverse classes of physiological triggers.", "doi": "10.1038/srep10274", "pmcid": "PMC4432564", "issn": "2045-2322", "publisher": "Nature Publishing Group", "publication": "Scientific Reports", "publication_date": "2015-05", "series_number": "5", "volume": "5", "issue": "5", "pages": "Art. No. 10274" }, { "id": "authors:1n749-wdv15", "collection": "authors", "collection_id": "1n749-wdv15", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20150409-093248232", "type": "article", "title": "Indigenous Bacteria from the Gut Microbiota Regulate Host Serotonin Biosynthesis", "author": [ { "family_name": "Yano", "given_name": "Jessica M.", "clpid": "Yano-Jessica-M" }, { "family_name": "Yu", "given_name": "Kristie", "orcid": "0000-0001-6735-3968", "clpid": "Yu-Kristie-B" }, { "family_name": "Donaldson", "given_name": "Gregory P.", "orcid": "0000-0002-8551-374X", "clpid": "Donaldson-Gregory-P" }, { "family_name": "Shastri", "given_name": "Gauri G.", "clpid": "Shastri-Gauri-G" }, { "family_name": "Ann", "given_name": "Phoebe", "clpid": "Ann-Phoebe" }, { "family_name": "Ma", "given_name": "Liang", "clpid": "Ma-Liang" }, { "family_name": "Nagler", "given_name": "Cathryn R.", "orcid": "0000-0001-7254-6617", "clpid": "Nagler-Cathryn-R" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Mazmanian", "given_name": "Sarkis K.", "orcid": "0000-0003-2713-1513", "clpid": "Mazmanian-S-K" }, { "family_name": "Hsiao", "given_name": "Elaine Y.", "orcid": "0000-0002-1633-588X", "clpid": "Hsiao-Elaine-Y-Bio" } ], "abstract": "The gastrointestinal (GI) tract contains much of the body's serotonin (5-hydroxytryptamine, 5-HT), but mechanisms controlling the metabolism of gut-derived 5-HT remain unclear. Here, we demonstrate that the microbiota plays a critical role in regulating host 5-HT. Indigenous spore-forming bacteria (Sp) from the mouse and human microbiota promote 5-HT biosynthesis from colonic enterochromaffin cells (ECs), which supply 5-HT to the mucosa, lumen, and circulating platelets. Importantly, microbiota-dependent effects on gut 5-HT significantly impact host physiology, modulating GI motility and platelet function. We identify select fecal metabolites that are increased by Sp and that elevate 5-HT in chromaffin cell cultures, suggesting direct metabolic signaling of gut microbes to ECs. Furthermore, elevating luminal concentrations of particular microbial metabolites increases colonic and blood 5-HT in germ-free mice. Altogether, these findings demonstrate that Sp are important modulators of host 5-HT and further highlight a key role for host-microbiota interactions in regulating fundamental 5-HT-related biological processes.", "doi": "10.1016/j.cell.2015.02.047", "pmcid": "PMC4393509", "issn": "0092-8674", "publisher": "Cell Press", "publication": "Cell", "publication_date": "2015-04-09", "series_number": "2", "volume": "161", "issue": "2", "pages": "264-276" }, { "id": "authors:3kz0k-z7430", "collection": "authors", "collection_id": "3kz0k-z7430", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20140925-085504154", "type": "article", "title": "The pumping lid: investigating multi-material 3D printing for equipment-free, programmable generation of positive and negative pressures for microfluidic applications", "author": [ { "family_name": "Begolo", "given_name": "Stefano", "clpid": "Begolo-S" }, { "family_name": "Zhukov", "given_name": "Dmitriy V.", "orcid": "0000-0002-4834-3147", "clpid": "Zhukov-D-V" }, { "family_name": "Selck", "given_name": "David A.", "clpid": "Selck-D-A" }, { "family_name": "Li", "given_name": "Liang", "clpid": "Li-Liang" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Equipment-free pumping is a challenging problem and an active area of research in microfluidics, with applications for both laboratory and limited-resource settings. This paper describes the pumping lid method, a strategy to achieve equipment-free pumping by controlled generation of pressure. Pressure was generated using portable, lightweight, and disposable parts that can be integrated with existing microfluidic devices to simplify workflow and eliminate the need for pumping equipment. The development of this method was enabled by multi-material 3D printing, which allows fast prototyping, including composite parts that combine materials with different mechanical properties (e.g. both rigid and elastic materials in the same part). The first type of pumping lid we describe was used to produce predictable positive or negative pressures via controlled compression or expansion of gases. A model was developed to describe the pressures and flow rates generated with this approach and it was validated experimentally. Pressures were pre-programmed by the geometry of the parts and could be tuned further even while the experiment was in progress. Using multiple lids or a composite lid with different inlets enabled several solutions to be pumped independently in a single device. The second type of pumping lid, which relied on vapor\u2013liquid equilibrium to generate pressure, was designed, modeled, and experimentally characterized. The pumping lid method was validated by controlling flow in different types of microfluidic applications, including the production of droplets, control of laminar flow profiles, and loading of SlipChip devices. We believe that applying the pumping lid methodology to existing microfluidic devices will enhance their use as portable diagnostic tools in limited resource settings as well as accelerate adoption of microfluidics in laboratories.", "doi": "10.1039/C4LC00910J", "pmcid": "PMC10773560", "issn": "1473-0197", "publisher": "Royal Society of Chemistry", "publication": "Lab on a Chip", "publication_date": "2014-12-21", "series_number": "24", "volume": "14", "issue": "24", "pages": "4616-4628" }, { "id": "authors:n8jjf-1v846", "collection": "authors", "collection_id": "n8jjf-1v846", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20141212-133134775", "type": "article", "title": "Advancing the speed, sensitivity and accuracy of biomolecular detection using multi-length-scale engineering", "author": [ { "family_name": "Kelley", "given_name": "Shana O.", "orcid": "0000-0003-3360-5359", "clpid": "Kelley-S-O" }, { "family_name": "Mirkin", "given_name": "Chad A.", "orcid": "0000-0002-6634-7627", "clpid": "Mirkin-C-A" }, { "family_name": "Walt", "given_name": "David R.", "clpid": "Walt-D-R" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Toner", "given_name": "Mehmet", "clpid": "Toner-M" }, { "family_name": "Sargent", "given_name": "Edward H.", "orcid": "0000-0003-0396-6495", "clpid": "Sargent-E-H" } ], "abstract": "Rapid progress in identifying disease biomarkers has increased the importance of creating high-performance detection technologies. Over the last decade, the design of many detection platforms has focused on either the nano or micro length scale. Here, we review recent strategies that combine nano- and microscale materials and devices to produce large improvements in detection sensitivity, speed and accuracy, allowing previously undetectable biomarkers to be identified in clinical samples. Microsensors that incorporate nanoscale features can now rapidly detect disease-related nucleic acids expressed in patient samples. New microdevices that separate large clinical samples into nanocompartments allow precise quantitation of analytes, and microfluidic systems that utilize nanoscale binding events can detect rare cancer cells in the bloodstream more accurately than before. These advances will lead to faster and more reliable clinical diagnostic devices.", "doi": "10.1038/NNANO.2014.261", "pmcid": "PMC4472305", "issn": "1748-3387", "publisher": "Nature Publishing Group", "publication": "Nature Nanotechnology", "publication_date": "2014-12", "series_number": "12", "volume": "9", "issue": "12", "pages": "969-980" }, { "id": "authors:v0dw7-bhr74", "collection": "authors", "collection_id": "v0dw7-bhr74", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20140829-120953993", "type": "article", "title": "Rapid fucosylation of intestinal epithelium sustains host\u2013commensal symbiosis in sickness", "author": [ { "family_name": "Pickard", "given_name": "Joseph M.", "clpid": "Pickard-J-M" }, { "family_name": "Maurice", "given_name": "Corinne F.", "clpid": "Maurice-C-F" }, { "family_name": "Kinnebrew", "given_name": "Melissa A.", "clpid": "Kinnebrew-M-A" }, { "family_name": "Abt", "given_name": "Michael C.", "clpid": "Abt-M-C" }, { "family_name": "Schenten", "given_name": "Diminik", "clpid": "Schenten-D" }, { "family_name": "Golovkina", "given_name": "Tatyana V.", "clpid": "Golovkina-T-V" }, { "family_name": "Bogatyrev", "given_name": "Said R.", "orcid": "0000-0003-0486-9451", "clpid": "Bogatyrev-S-R" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Pamer", "given_name": "Eric G.", "clpid": "Pamer-E-G" }, { "family_name": "Turnbaugh", "given_name": "Peter J.", "clpid": "Turnbaugh-P-J" }, { "family_name": "Chrenovsky", "given_name": "Alexander V.", "clpid": "Chernovsky-A-V" } ], "abstract": "Systemic infection induces conserved physiological responses that include both resistance and 'tolerance of infection' mechanisms. Temporary anorexia associated with an infection is often beneficial, reallocating energy from food foraging towards resistance to infection or depriving pathogens of nutrients. However, it imposes a stress on intestinal commensals, as they also experience reduced substrate availability; this affects host fitness owing to the loss of caloric intake and colonization resistance (protection from additional infections). We hypothesized that the host might utilize internal resources to support the gut microbiota during the acute phase of the disease. Here we show that systemic exposure to Toll-like receptor (TLR) ligands causes rapid \u03b1(1,2)-fucosylation of small intestine epithelial cells (IECs) in mice, which requires the sensing of TLR agonists, as well as the production of interleukin (IL)-23 by dendritic cells, activation of innate lymphoid cells and expression of fucosyltransferase 2 (Fut2) by IL-22-stimulated IECs. Fucosylated proteins are shed into the lumen and fucose is liberated and metabolized by the gut microbiota, as shown by reporter bacteria and community-wide analysis of microbial gene expression. Fucose affects the expression of microbial metabolic pathways and reduces the expression of bacterial virulence genes. It also improves host tolerance of the mild pathogen Citrobacter rodentium. Thus, rapid IEC fucosylation appears to be a protective mechanism that utilizes the host's resources to maintain host\u2013microbial interactions during pathogen-induced stress.", "doi": "10.1038/nature13823", "pmcid": "PMC4214913", "issn": "0028-0836", "publisher": "Nature Publishing Group", "publication": "Nature", "publication_date": "2014-10-30", "series_number": "7524", "volume": "514", "issue": "7524", "pages": "638-641" }, { "id": "authors:fxrae-42803", "collection": "authors", "collection_id": "fxrae-42803", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20141015-105804358", "type": "article", "title": "Digital, Ultrasensitive, End-Point Protein Measurements with Large Dynamic Range via Brownian Trapping with Drift", "author": [ { "family_name": "Ge", "given_name": "Shencheng", "clpid": "Ge-Shencheng" }, { "family_name": "Liu", "given_name": "Weishan", "clpid": "Liu-Weishan" }, { "family_name": "Schlappi", "given_name": "Travis", "orcid": "0000-0001-6132-6459", "clpid": "Schlappi-T-S" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This communication shows that the concept of Brownian trapping with drift can be applied to improve quantitative molecular measurements. It has the potential to combine the robustness of end-point spatially resolved readouts, the ultrasensitivity of digital single-molecule measurements, and the large dynamic range of qPCR; furthermore, at low concentrations of analytes, it can provide a direct comparison of the signals arising from the analyte and from the background. It relies on the finding that molecules simultaneously diffusing, drifting (via slow flow), and binding to an array of nonsaturable surface traps have an exponentially decreasing probability of escaping the traps over time and therefore give rise to an exponentially decaying distribution of trapped molecules in space. This concept was tested with enzyme and protein measurements in a microfluidic device.", "doi": "10.1021/ja507849b", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2014-10-22", "series_number": "42", "volume": "136", "issue": "42", "pages": "14662-14665" }, { "id": "authors:y9fxh-aht69", "collection": "authors", "collection_id": "y9fxh-aht69", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20140606-091339805", "type": "article", "title": "Digital biology and chemistry", "author": [ { "family_name": "Witters", "given_name": "Daan", "clpid": "Witters-D" }, { "family_name": "Sun", "given_name": "Bing", "clpid": "Sun-Bing" }, { "family_name": "Begolo", "given_name": "Stefano", "clpid": "Begolo-S" }, { "family_name": "Rodriguez-Manzano", "given_name": "Jesus", "clpid": "Rodriguez-Manzano-J" }, { "family_name": "Robles", "given_name": "Whitney", "clpid": "Robles-W" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This account examines developments in \"digital\" biology and chemistry within the context of microfluidics, from a personal perspective. Using microfluidics as a frame of reference, we identify two areas of research within digital biology and chemistry that are of special interest: (i) the study of systems that switch between discrete states in response to changes in chemical concentration of signals, and (ii) the study of single biological entities such as molecules or cells. In particular, microfluidics accelerates analysis of switching systems (i.e., those that exhibit a sharp change in output over a narrow range of input) by enabling monitoring of multiple reactions in parallel over a range of concentrations of signals. Conversely, such switching systems can be used to create new kinds of microfluidic detection systems that provide \"analog-to-digital\" signal conversion and logic. Microfluidic compartmentalization technologies for studying and isolating single entities can be used to reconstruct and understand cellular processes, study interactions between single biological entities, and examine the intrinsic heterogeneity of populations of molecules, cells, or organisms. Furthermore, compartmentalization of single cells or molecules in \"digital\" microfluidic experiments can induce switching in a range of reaction systems to enable sensitive detection of cells or biomolecules, such as with digital ELISA or digital PCR. This \"digitizing\" offers advantages in terms of robustness, assay design, and simplicity because quantitative information can be obtained with qualitative measurements. While digital formats have been shown to improve the robustness of existing chemistries, we anticipate that in the future they will enable new chemistries to be used for quantitative measurements, and that digital biology and chemistry will continue to provide further opportunities for measuring biomolecules, understanding natural systems more deeply, and advancing molecular and cellular analysis. Microfluidics will impact digital biology and chemistry and will also benefit from them if it becomes massively distributed.", "doi": "10.1039/c4lc00248b", "issn": "1473-0197", "publisher": "Royal Society of Chemistry", "publication": "Lab on a Chip", "publication_date": "2014-09-07", "series_number": "17", "volume": "14", "issue": "17", "pages": "3225-3232" }, { "id": "authors:72rme-7f619", "collection": "authors", "collection_id": "72rme-7f619", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20140620-180156638", "type": "article", "title": "Individually addressable arrays of replica microbial cultures enabled by splitting SlipChips", "author": [ { "family_name": "Ma", "given_name": "Liang", "clpid": "Ma-Liang" }, { "family_name": "Datta", "given_name": "Sujit S.", "orcid": "0000-0003-2400-1561", "clpid": "Datta-S-S" }, { "family_name": "Karymov", "given_name": "Mikhail A.", "clpid": "Karymov-M-A" }, { "family_name": "Pan", "given_name": "Qichao", "clpid": "Pan-Qichao" }, { "family_name": "Begolo", "given_name": "Stefano", "clpid": "Begolo-S" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Isolating microbes carrying genes of interest from environmental samples is important for applications in biology and medicine. However, this involves the use of genetic assays that often require lysis of microbial cells, which is not compatible with the goal of obtaining live cells for isolation and culture. This paper describes the design, fabrication, biological validation, and underlying physics of a microfluidic SlipChip device that addresses this challenge. The device is composed of two conjoined plates containing 1000 microcompartments, each comprising two juxtaposed wells, one on each opposing plate. Single microbial cells are stochastically confined and subsequently cultured within the microcompartments. Then, we split each microcompartment into two replica droplets, both containing microbial culture, and then controllably separate the two plates while retaining each droplet within each well. We experimentally describe the droplet retention as a function of capillary pressure, viscous pressure, and viscosity of the aqueous phase. Within each pair of replicas, one can be used for genetic analysis, and the other preserves live cells for growth. This microfluidic approach provides a facile way to cultivate anaerobes from complex communities. We validate this method by targeting, isolating, and culturing Bacteroides vulgatus, a core gut anaerobe, from a clinical sample. To date, this methodology has enabled isolation of a novel microbial taxon, representing a new genus. This approach could also be extended to the study of other microorganisms and even mammalian systems, and may enable targeted retrieval of solutions in applications including digital PCR, sequencing, single cell analysis, and protein crystallization.", "doi": "10.1039/c4ib00109e", "pmcid": "PMC4131746", "issn": "1757-9694", "publisher": "Royal Society of Chemistry", "publication": "Integrative Biology", "publication_date": "2014-08", "series_number": "8", "volume": "6", "issue": "8", "pages": "796-805" }, { "id": "authors:zz27g-zjb70", "collection": "authors", "collection_id": "zz27g-zjb70", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20140609-150635938", "type": "article", "title": "Measuring Fate and Rate of Single-Molecule Competition of Amplification and Restriction Digestion, and Its Use for Rapid Genotyping Tested with Hepatitis C Viral RNA", "author": [ { "family_name": "Sun", "given_name": "Bing", "clpid": "Sun-Bing" }, { "family_name": "Rodriguez-Manzano", "given_name": "Jesus", "clpid": "Rodriguez-Manzano-Jesus" }, { "family_name": "Selck", "given_name": "David A.", "clpid": "Selck-David-A" }, { "family_name": "Khorosheva", "given_name": "Eugenia", "clpid": "Korosheva-Eugenia" }, { "family_name": "Karymov", "given_name": "Mikhail A.", "clpid": "Karymov-Mikhail-A" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "We experimentally monitored, at the single-molecule level, the competition among reverse transcription, exponential amplification (RT-LAMP), and linear degradation (restriction enzymes) starting with hepatitis C viral RNA molecules. We found significant heterogeneity in the rate of single-molecule amplification; introduction of the restriction enzymes affected both the rate and the \"fate\" (the binary outcome) of single-molecule amplification. While end-point digital measurements were primarily sensitive to changes in fate, the bulk real-time kinetic measurements were dominated by the rate of amplification of the earliest molecules, and were not sensitive to fate of the rest of the molecules. We show how this competition of reactions can be used for rapid HCV genotyping with either digital or bulk readout. This work advances our understanding of single-molecule dynamics in reaction networks and may help bring genotyping capabilities out of clinical labs and into limited-resource settings.", "doi": "10.1002/ange.201403035", "pmcid": "PMC4116457", "issn": "1433-7851", "publisher": "Wiley", "publication": "Angewandte Chemie International Edition", "publication_date": "2014-07-28", "series_number": "31", "volume": "53", "issue": "31", "pages": "8088-8092" }, { "id": "authors:1t9mb-4k126", "collection": "authors", "collection_id": "1t9mb-4k126", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20140620-103511347", "type": "article", "title": "Gene-targeted microfluidic cultivation validated by isolation of a gut bacterium listed in Human Microbiome Project's Most Wanted taxa", "author": [ { "family_name": "Ma", "given_name": "Liang", "clpid": "Ma-Liang" }, { "family_name": "Kim", "given_name": "Jungwoo", "orcid": "0000-0002-5215-2044", "clpid": "Kim-Jungwoo" }, { "family_name": "Hatzenpichler", "given_name": "Roland", "orcid": "0000-0002-5489-3444", "clpid": "Hatzenpichler-R" }, { "family_name": "Karymov", "given_name": "Mikhail A.", "clpid": "Karymov-M-A" }, { "family_name": "Hubert", "given_name": "Nathaniel", "clpid": "Hubert-N" }, { "family_name": "Hanan", "given_name": "Ira M.", "clpid": "Hanan-I-M" }, { "family_name": "Chang", "given_name": "Eugene B.", "clpid": "Chang-Eugene-B" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper describes a microfluidics-based workflow for genetically targeted isolation and cultivation of microorganisms from complex clinical samples. Data sets from high-throughput sequencing suggest the existence of previously unidentified bacterial taxa and functional genes with high biomedical importance. Obtaining isolates of these targets, preferably in pure cultures, is crucial for advancing understanding of microbial genetics and physiology and enabling physical access to microbes for further applications. However, the majority of microbes have not been cultured, due in part to the difficulties of both identifying proper growth conditions and characterizing and isolating each species. We describe a method that enables genetically targeted cultivation of microorganisms through a combination of microfluidics and on- and off-chip assays. This method involves (i) identification of cultivation conditions for microbes using growth substrates available only in small quantities as well as the correction of sampling bias using a \"chip wash\" technique; and (ii) performing on-chip genetic assays while also preserving live bacterial cells for subsequent scale-up cultivation of desired microbes, by applying recently developed technology to create arrays of individually addressable replica microbial cultures. We validated this targeted approach by cultivating a bacterium, here referred to as isolate microfluidicus 1, from a human cecal biopsy. Isolate microfluidicus 1 is, to our knowledge, the first successful example of targeted cultivation of a microorganism from the high-priority group of the Human Microbiome Project's \"Most Wanted\" list, and, to our knowledge, the first cultured representative of a previously unidentified genus of the Ruminococcaceae family.", "doi": "10.1073/pnas.1404753111", "pmcid": "PMC4103313", "issn": "0027-8424", "publisher": "National Academy of Sciences", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "publication_date": "2014-07-08", "series_number": "27", "volume": "111", "issue": "27", "pages": "9768-9773" }, { "id": "authors:bhj62-sf283", "collection": "authors", "collection_id": "bhj62-sf283", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20131108-122433726", "type": "article", "title": "Increased Robustness of Single-Molecule Counting with Microfluidics, Digital Isothermal Amplification, and a Mobile Phone versus Real-Time Kinetic Measurements", "author": [ { "family_name": "Selck", "given_name": "David A.", "clpid": "Selck-D-A" }, { "family_name": "Karymov", "given_name": "Mikhail A.", "clpid": "Karymov-M-A" }, { "family_name": "Sun", "given_name": "Bing", "clpid": "Sun-Bing" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Quantitative bioanalytical measurements are commonly performed in a kinetic format and are known to not be robust to perturbation that affects the kinetics itself or the measurement of kinetics. We hypothesized that the same measurements performed in a \"digital\" (single-molecule) format would show increased robustness to such perturbations. Here, we investigated the robustness of an amplification reaction (reverse-transcription loop-mediated amplification, RT-LAMP) in the context of fluctuations in temperature and time when this reaction is used for quantitative measurements of HIV-1 RNA molecules under limited-resource settings (LRS). The digital format that counts molecules using dRT-LAMP chemistry detected a 2-fold change in concentration of HIV-1 RNA despite a 6 \u00b0C temperature variation (p-value = 6.7 \u00d7 10^\u20137), whereas the traditional kinetic (real-time) format did not (p-value = 0.25). Digital analysis was also robust to a 20 min change in reaction time, to poor imaging conditions obtained with a consumer cell-phone camera, and to automated cloud-based processing of these images (R^2 = 0.9997 vs true counts over a 100-fold dynamic range). Fluorescent output of multiplexed PCR amplification could also be imaged with the cell phone camera using flash as the excitation source. Many nonlinear amplification schemes based on organic, inorganic, and biochemical reactions have been developed, but their robustness is not well understood. This work implies that these chemistries may be significantly more robust in the digital, rather than kinetic, format. It also calls for theoretical studies to predict robustness of these chemistries and, more generally, to design robust reaction architectures. The SlipChip that we used here and other digital microfluidic technologies already exist to enable testing of these predictions. Such work may lead to identification or creation of robust amplification chemistries that enable rapid and precise quantitative molecular measurements under LRS. Furthermore, it may provide more general principles describing robustness of chemical and biological networks in digital formats.", "doi": "10.1021/ac4030413", "pmcid": "PMC3924768", "issn": "0003-2700", "publisher": "American Chemical Society", "publication": "Analytical Chemistry", "publication_date": "2013-11-07", "series_number": "22", "volume": "85", "issue": "22", "pages": "11129-11136" }, { "id": "authors:5pedx-y1p08", "collection": "authors", "collection_id": "5pedx-y1p08", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20131024-155658108", "type": "article", "title": "Chemical Analog-to-Digital Signal Conversion Based on Robust Threshold Chemistry and Its Evaluation in the Context of Microfluidics-Based Quantitative Assays", "author": [ { "family_name": "Huynh", "given_name": "Toan", "clpid": "Huynh-Toan" }, { "family_name": "Sun", "given_name": "Bing", "clpid": "Sun-Bing" }, { "family_name": "Li", "given_name": "Liang", "clpid": "Li-Liang" }, { "family_name": "Nichols", "given_name": "Kevin P.", "clpid": "Nichols-K-P" }, { "family_name": "Koyner", "given_name": "Jay L.", "clpid": "Koyner-J-L" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "In this article, we describe a nonlinear threshold chemistry based on enzymatic inhibition and demonstrate how it can be coupled with microfluidics to convert a chemical concentration (analog input) into patterns of ON or OFF reaction outcomes (chemical digital readout). Quantification of small changes in concentration is needed in a number of assays, such as that for cystatin C, where a 1.5-fold increase in concentration may indicate the presence of acute kidney injury or progression of chronic kidney disease. We developed an analog-to-digital chemical signal conversion that gives visual readout and applied it to an assay for cystatin C as a model target. The threshold chemistry is based on enzymatic inhibition and gives sharper responses with tighter inhibition. The chemistry described here uses acetylcholinesterase (AChE) and produces an unambiguous color change when the input is above a predetermined threshold concentration. An input gives a pattern of ON/OFF responses when subjected to a monotonic sequence of threshold concentrations, revealing the input concentration at the point of transition from OFF to ON outcomes. We demonstrated that this threshold chemistry can detect a 1.30-fold increase in concentration at 22 \u00b0C and that it is robust to experimental fluctuations: it provided the same output despite changes in temperature (22\u201334 \u00b0C) and readout time (10-fold range). We applied this threshold chemistry to diagnostics by coupling it with a traditional sandwich immunoassay for serum cystatin C. Because one quantitative measurement comprises several assays, each with its own threshold concentration, we used a microfluidic SlipChip device to process 12 assays in parallel, detecting a 1.5-fold increase (from 0.64 (49 nM) to 0.96 mg/L (74 nM)) of cystatin C in serum. We also demonstrated applicability to analysis of patient serum samples and the ability to image results using a cell phone camera. This work indicates that combining developments in nonlinear chemistries with microfluidics may lead to development of user-friendly diagnostic assays with simple readouts.", "doi": "10.1021/ja4062882", "pmcid": "PMC3860884", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2013-10-02", "series_number": "39", "volume": "135", "issue": "39", "pages": "14775-14783" }, { "id": "authors:ynbt6-p8710", "collection": "authors", "collection_id": "ynbt6-p8710", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20131002-113705785", "type": "article", "title": "A microfluidic device for dry sample preservation in remote settings", "author": [ { "family_name": "Begolo", "given_name": "Stefano", "clpid": "Begolo-S" }, { "family_name": "Shen", "given_name": "Feng", "orcid": "0000-0002-4709-330X", "clpid": "Shen-Feng" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper describes a microfluidic device for dry preservation of biological specimens at room temperature that\nincorporates chemical stabilization matrices. Long-term stabilization of samples is crucial for remote medical analysis,\nbiosurveillance, and archiving, but the current paradigm for transporting remotely obtained samples relies on\nthe costly \"cold chain\" to preserve analytes within biospecimens. We propose an alternative approach that involves\nthe use of microfluidics to preserve samples in the dry state with stabilization matrices, developed by others, that\nare based on self-preservation chemistries found in nature. We describe a SlipChip-based device that allows minimally\ntrained users to preserve samples with the three simple steps of placing a sample at an inlet, closing a lid,\nand slipping one layer of the device. The device fills automatically, and a pre-loaded desiccant dries the samples.\nLater, specimens can be rehydrated and recovered for analysis in a laboratory. This device is portable, compact,\nand self-contained, so it can be transported and operated by untrained users even in limited-resource settings. Features\nsuch as dead-end and sequential filling, combined with a \"pumping lid\" mechanism, enable precise quantification\nof the original sample's volume while avoiding overfilling. In addition, we demonstrated that the device can\nbe integrated with a plasma filtration module, and we validated device operations and capabilities by testing the\nstability of purified RNA solutions. These features and the modularity of this platform (which facilitates integration\nand simplifies operation) would be applicable to other microfluidic devices beyond this application. We envision\nthat as the field of stabilization matrices develops, microfluidic devices will be useful for cost-effectively facilitating\nremote analysis and biosurveillance while also opening new opportunities for diagnostics, drug development, and\nother medical fields.", "doi": "10.1039/c3lc50747e", "pmcid": "PMC3851311", "issn": "1473-0197", "publisher": "Royal Society of Chemistry", "publication": "Lab on a Chip", "publication_date": "2013-09-17", "series_number": "22", "volume": "13", "issue": "22", "pages": "4331-4342" }, { "id": "authors:rdfze-ycn24", "collection": "authors", "collection_id": "rdfze-ycn24", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130404-103625840", "type": "article", "title": "A Genetically Encoded AND Gate for Cell-Targeted Metabolic Labeling of Proteins", "author": [ { "family_name": "Mahdavi", "given_name": "Alborz", "orcid": "0000-0002-8790-8112", "clpid": "Mahdavi-Alborz" }, { "family_name": "Segall-Shapiro", "given_name": "Thomas H.", "clpid": "Segall-Shapiro-T-H" }, { "family_name": "Kou", "given_name": "Songzi", "clpid": "Kou-Songzi" }, { "family_name": "Jindal", "given_name": "Granton A.", "clpid": "Jindal-G-A" }, { "family_name": "Hoff", "given_name": "Kevin G.", "clpid": "Hoff-K-G" }, { "family_name": "Liu", "given_name": "Shirley", "clpid": "Liu-Shirley" }, { "family_name": "Chitsaz", "given_name": "Mohsen", "clpid": "Chitsaz-M" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Silberg", "given_name": "Jonathan J.", "clpid": "Silberg-J-J" }, { "family_name": "Tirrell", "given_name": "David A.", "orcid": "0000-0003-3175-4596", "clpid": "Tirrell-D-A" } ], "abstract": "We describe a genetic AND gate for cell-targeted metabolic labeling and proteomic analysis in complex cellular systems. The centerpiece of the AND gate is a bisected methionyl-tRNA synthetase (MetRS) that charges the Met surrogate azidonorleucine (Anl) to tRNAMet. Cellular protein labeling occurs only upon activation of two different promoters that drive expression of the N- and C-terminal fragments of the bisected MetRS. Anl-labeled proteins can be tagged with fluorescent dyes or affinity reagents via either copper-catalyzed or strain-promoted azide\u2013alkyne cycloaddition. Protein labeling is apparent within 5 min after addition of Anl to bacterial cells in which the AND gate has been activated. This method allows spatial and temporal control of proteomic labeling and identification of proteins made in specific cellular subpopulations. The approach is demonstrated by selective labeling of proteins in bacterial cells immobilized in the center of a laminar-flow microfluidic channel, where they are exposed to overlapping, opposed gradients of inducers of the N- and C-terminal MetRS fragments. The observed labeling profile is predicted accurately from the strengths of the individual input signals.", "doi": "10.1021/ja400448f", "pmcid": "PMC3620012", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2013-02-27", "series_number": "8", "volume": "135", "issue": "8", "pages": "2979-2982" }, { "id": "authors:ye8xy-tn052", "collection": "authors", "collection_id": "ye8xy-tn052", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130320-140730972", "type": "article", "title": "Mechanistic Evaluation of the Pros and Cons of Digital RT-LAMP for HIV-1 Viral Load Quantification on a Microfluidic Device and Improved Efficiency via a Two-Step Digital Protocol", "author": [ { "family_name": "Sun", "given_name": "Bing", "clpid": "Sun-Bing" }, { "family_name": "Shen", "given_name": "Feng", "orcid": "0000-0002-4709-330X", "clpid": "Shen-Feng" }, { "family_name": "McCalla", "given_name": "Stephanie E.", "clpid": "McCalla-S-E" }, { "family_name": "Kreutz", "given_name": "Jason E.", "clpid": "Kreutz-J-E" }, { "family_name": "Karymov", "given_name": "Mikhail A.", "clpid": "Karymov-M-A" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Here we used a SlipChip microfluidic device to evaluate the performance of digital reverse transcription-loop-mediated isothermal amplification (dRT-LAMP) for quantification of HIV viral RNA. Tests are needed for monitoring HIV viral load to control the emergence of drug resistance and to diagnose acute HIV infections. In resource-limited settings, in vitro measurement of HIV viral load in a simple format is especially needed, and single-molecule counting using a digital format could provide a potential solution. We showed here that when one-step dRT-LAMP is used for quantification of HIV RNA, the digital count is lower than expected and is limited by the yield of desired cDNA. We were able to overcome the limitations by developing a microfluidic protocol to manipulate many single molecules in parallel through a two-step digital process. In the first step we compartmentalize the individual RNA molecules (based on Poisson statistics) and perform reverse transcription on each RNA molecule independently to produce DNA. In the second step, we perform the LAMP amplification on all individual DNA molecules in parallel. Using this new protocol, we increased the absolute efficiency (the ratio between the concentration calculated from the actual count and the expected concentration) of dRT-LAMP 10-fold, from 2% to 23%, by (i) using a more efficient reverse transcriptase, (ii) introducing RNase H to break up the DNA:RNA hybrid, and (iii) adding only the BIP primer during the RT step. We also used this two-step method to quantify HIV RNA purified from four patient samples and found that in some cases, the quantification results were highly sensitive to the sequence of the patient's HIV RNA. We learned the following three lessons from this work: (i) digital amplification technologies, including dLAMP and dPCR, may give adequate dilution curves and yet have low efficiency, thereby providing quantification values that underestimate the true concentration. Careful validation is essential before a method is considered to provide absolute quantification; (ii) the sensitivity of dLAMP to the sequence of the target nucleic acid necessitates additional validation with patient samples carrying the full spectrum of mutations; (iii) for multistep digital amplification chemistries, such as a combination of reverse transcription with amplification, microfluidic devices may be used to decouple these steps from one another and to perform them under different, individually optimized conditions for improved efficiency.", "doi": "10.1021/ac3037206", "pmcid": "PMC3578705", "issn": "0003-2700", "publisher": "American Chemical Society", "publication": "Analytical Chemistry", "publication_date": "2013-02-05", "series_number": "3", "volume": "85", "issue": "3", "pages": "1540-1546" }, { "id": "authors:kjj6n-4nc93", "collection": "authors", "collection_id": "kjj6n-4nc93", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20120222-071637506", "type": "article", "title": "Control of Initiation, Rate, and Routing of Spontaneous Capillary-Driven Flow of Liquid Droplets through Microfluidic Channels on SlipChip", "author": [ { "family_name": "Pompano", "given_name": "Rebecca R.", "clpid": "Pompano-R-R" }, { "family_name": "Platt", "given_name": "Carol E.", "clpid": "Platt-C-E" }, { "family_name": "Karymov", "given_name": "Mikhail A.", "clpid": "Karymov-M-A" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This Article describes the use of capillary pressure to initiate and control the rate of spontaneous liquid\u2013liquid flow through microfluidic channels. In contrast to flow driven by external pressure, flow driven by capillary pressure is dominated by interfacial phenomena and is exquisitely sensitive to the chemical composition and geometry of the fluids and channels. A stepwise change in capillary force was initiated on a hydrophobic SlipChip by slipping a shallow channel containing an aqueous droplet into contact with a slightly deeper channel filled with immiscible oil. This action induced spontaneous flow of the droplet into the deeper channel. A model predicting the rate of spontaneous flow was developed on the basis of the balance of net capillary force with viscous flow resistance, using as inputs the liquid\u2013liquid surface tension, the advancing and receding contact angles at the three-phase aqueous\u2013oil\u2013surface contact line, and the geometry of the devices. The impact of contact angle hysteresis, the presence or absence of a lubricating oil layer, and adsorption of surface-active compounds at liquid\u2013liquid or liquid\u2013solid interfaces were quantified. Two regimes of flow spanning a 104-fold range of flow rates were obtained and modeled quantitatively, with faster (mm/s) flow obtained when oil could escape through connected channels as it was displaced by flowing aqueous solution, and slower (micrometer/s) flow obtained when oil escape was mostly restricted to a micrometer-scale gap between the plates of the SlipChip (\"dead-end flow\"). Rupture of the lubricating oil layer (reminiscent of a Cassie\u2013Wenzel transition) was proposed as a cause of discrepancy between the model and the experiment. Both dilute salt solutions and complex biological solutions such as human blood plasma could be flowed using this approach. We anticipate that flow driven by capillary pressure will be useful for the design and operation of flow in microfluidic applications that do not require external power, valves, or pumps, including on SlipChip and other droplet- or plug-based microfluidic devices. In addition, this approach may be used as a sensitive method of evaluating interfacial tension, contact angles, and wetting phenomena on chip.", "doi": "10.1021/la204399m", "pmcid": "PMC3271727", "issn": "0743-7463", "publisher": "American Chemical Society", "publication": "Langmuir", "publication_date": "2012-01-24", "series_number": "3", "volume": "28", "issue": "3", "pages": "1931-1941" }, { "id": "authors:ytsqa-axv28", "collection": "authors", "collection_id": "ytsqa-axv28", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20111116-090435392", "type": "article", "title": "Multiplexed Quantification of Nucleic Acids with Large Dynamic\n Range Using Multivolume Digital RT-PCR on a Rotational SlipChip Tested with HIV and Hepatitis C Viral Load", "author": [ { "family_name": "Shen", "given_name": "Feng", "orcid": "0000-0002-4709-330X", "clpid": "Shen-Feng" }, { "family_name": "Sun", "given_name": "Bing", "clpid": "Sun-Bing" }, { "family_name": "Kreutz", "given_name": "Jason E.", "clpid": "Kreutz-J-E" }, { "family_name": "Davydova", "given_name": "Elena K.", "clpid": "Davydova-E-K" }, { "family_name": "Du", "given_name": "Wenbin", "orcid": "0000-0002-7401-1410", "clpid": "Du-Wenbin" }, { "family_name": "Reddy", "given_name": "Poluru L.", "clpid": "Reddy-P-L" }, { "family_name": "Joseph", "given_name": "Loren J.", "clpid": "Joseph-L-J" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "In this paper, we are working toward a problem of great importance to global health: determination of viral HIV and hepatitis C (HCV) loads under point-of-care and resource limited settings. While antiretroviral treatments are becoming widely available, viral load must be evaluated at regular intervals to prevent the spread of drug resistance and requires a quantitative measurement of RNA concentration over a wide dynamic range (from 50 up to 10^6 molecules/mL for HIV and up to 108 molecules/mL for HCV). \"Digital\" single molecule measurements are attractive for quantification, but the dynamic range of such systems is typically limited or requires excessive numbers of compartments. Here we designed and tested two microfluidic rotational SlipChips to perform multivolume digital RT-PCR (MV digital RT-PCR) experiments with large and tunable dynamic range. These designs were characterized using synthetic control RNA and validated with HIV viral RNA and HCV control viral RNA. The first design contained 160 wells of each of four volumes (125 nL, 25 nL, 5 nL, and 1 nL) to achieve a dynamic range of 5.2 \u00d7 10^2 to 4.0 \u00d7 10^6 molecules/mL at 3-fold resolution. The second design tested the flexibility of this approach, and further expanded it to allow for multiplexing while maintaining a large dynamic range by adding additional wells with volumes of 0.2 nL and 625 nL and dividing the SlipChip into five regions to analyze five samples each at a dynamic range of 1.8 \u00d7 10^3 to 1.2 \u00d7 10^7 molecules/mL at 3-fold resolution. No evidence of cross-contamination was observed. The multiplexed SlipChip can be used to analyze a single sample at a dynamic range of 1.7 \u00d7 10^2 to 2.0 \u00d7 10^7 molecules/mL at 3-fold resolution with limit of detection of 40 molecules/mL. HIV viral RNA purified from clinical samples were tested on the SlipChip, and viral load results were self-consistent and in good agreement with results determined using the Roche COBAS AmpliPrep/COBAS TaqMan HIV-1 Test. With further validation, this SlipChip should become useful to precisely quantify viral HIV and HCV RNA for high-performance diagnostics in resource-limited settings. These microfluidic designs should also be valuable for other diagnostic and research applications, including detecting rare cells and rare mutations, prenatal diagnostics, monitoring residual disease, and quantifying copy number variation and gene expression patterns. The theory for the design and analysis of multivolume digital PCR experiments is presented in other work by Kreutz et al.", "doi": "10.1021/ja2060116", "pmcid": "PMC3216675", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2011-11-09", "series_number": "44", "volume": "133", "issue": "44", "pages": "17705-17712" }, { "id": "authors:d3cnh-s9x12", "collection": "authors", "collection_id": "d3cnh-s9x12", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20111213-144640436", "type": "article", "title": "Theoretical Design and Analysis of Multivolume Digital Assays with Wide Dynamic Range Validated Experimentally with Microfluidic Digital PCR", "author": [ { "family_name": "Kreutz", "given_name": "Jason E.", "clpid": "Kreutz-J-E" }, { "family_name": "Munson", "given_name": "Todd", "clpid": "Munson-T" }, { "family_name": "Huynh", "given_name": "Toan", "clpid": "Huynh-Toan" }, { "family_name": "Shen", "given_name": "Feng", "orcid": "0000-0002-4709-330X", "clpid": "Shen-Feng" }, { "family_name": "Du", "given_name": "Wenbin", "orcid": "0000-0002-7401-1410", "clpid": "Du-Wenbin" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper presents a protocol using theoretical methods and free software to design and analyze multivolume digital PCR (MV digital PCR) devices; the theory and software are also applicable to design and analysis of dilution series in digital PCR. MV digital PCR minimizes the total number of wells required for \"digital\" (single molecule) measurements while maintaining high dynamic range and high resolution. In some examples, multivolume designs with fewer than 200 total wells are predicted to provide dynamic range with 5-fold resolution similar to that of single-volume designs requiring 12\u2009000 wells. Mathematical techniques were utilized and expanded to maximize the information obtained from each experiment and to quantify performance of devices and were experimentally validated using the SlipChip platform. MV digital PCR was demonstrated to perform reliably, and results from wells of different volumes agreed with one another. No artifacts due to different surface-to-volume ratios were observed, and single molecule amplification in volumes ranging from 1 to 125 nL was self-consistent. The device presented here was designed to meet the testing requirements for measuring clinically relevant levels of HIV viral load at the point-of-care (in plasma, <500 molecules/mL to >1\u2009000\u2009000 molecules/mL), and the predicted resolution and dynamic range was experimentally validated using a control sequence of DNA. This approach simplifies digital PCR experiments, saves space, and thus enables multiplexing using separate areas for each sample on one chip, and facilitates the development of new high-performance diagnostic tools for resource-limited applications. The theory and software presented here are general and are applicable to designing and analyzing other digital analytical platforms including digital immunoassays and digital bacterial analysis. It is not limited to SlipChip and could also be useful for the design of systems on platforms including valve-based and droplet-based platforms. In a separate publication by Shen et al. (J. Am. Chem. Soc., 2011, DOI: 10.1021/ja2060116), this approach is used to design and test digital RT-PCR devices for quantifying RNA.", "doi": "10.1021/ac201658s", "pmcid": "PMC3216679", "issn": "0003-2700", "publisher": "American Chemical Society", "publication": "Analytical Chemistry", "publication_date": "2011-11-01", "series_number": "21", "volume": "83", "issue": "21", "pages": "8158-8168" }, { "id": "authors:2ng39-t2e84", "collection": "authors", "collection_id": "2ng39-t2e84", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160728693", "type": "article", "title": "Toward Mechanistic Understanding of Nuclear Reprocessing Chemistries by Quantifying Lanthanide Solvent Extraction Kinetics via Microfluidics with Constant Interfacial Area and Rapid Mixing", "author": [ { "family_name": "Nichols", "given_name": "Kevin P.", "clpid": "Nichols-K-P" }, { "family_name": "Pompano", "given_name": "Rebecca R.", "clpid": "Pompano-R-R" }, { "family_name": "Li", "given_name": "Liang", "clpid": "Li-Liang" }, { "family_name": "Gelis", "given_name": "Artem V.", "clpid": "Gelis-A-V" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "The closing of the nuclear fuel cycle is an unsolved problem of great importance. Separating radionuclides produced in a nuclear reactor is useful both for the storage of nuclear waste and for recycling of nuclear fuel. These separations can be performed by designing appropriate chelation chemistries and liquid-liquid extraction schemes, such as in the TALSPEAK process (Trivalent Actinide-Lanthanide Separation by Phosphorus reagent Extraction from Aqueous Komplexes). However, there are no approved methods for the industrial scale reprocessing of civilian nuclear fuel in the United States. One bottleneck in the design of next-generation solvent extraction-based nuclear fuel reprocessing schemes is a lack of interfacial mass transfer rate constants obtained under well-controlled conditions for lanthanide and actinide ligand complexes; such rate constants are a prerequisite for mechanistic understanding of the extraction chemistries involved and are of great assistance in the design of new chemistries. In addition, rate constants obtained under conditions of known interfacial area have immediate, practical utility in models required for the scaling-up of laboratory-scale demonstrations to industrial-scale solutions. Existing experimental techniques for determining these rate constants suffer from two key drawbacks: either slow mixing or unknown interfacial area. The volume of waste produced by traditional methods is an additional, practical concern in experiments involving radioactive elements, both from disposal cost and experimenter safety standpoints. In this paper, we test a plug-based microfluidic system that uses flowing plugs (droplets) in microfluidic channels to determine absolute interfacial mass transfer rate constants under conditions of both rapid mixing and controlled interfacial area. We utilize this system to determine, for the first time, the rate constants for interfacial transfer of all lanthanides, minus promethium, plus yttrium, under TALSPEAK process conditions, as a first step toward testing the molecular mechanism of this separation process.", "doi": "10.1021/ja206020u", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2011-10-05", "series_number": "39", "volume": "133", "issue": "39", "pages": "15721-9" }, { "id": "authors:fgpqe-ff172", "collection": "authors", "collection_id": "fgpqe-ff172", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160716001", "type": "article", "title": "Three-Dimensional Nanocrystal Superlattices Grown in Nanoliter Microfluidic Plugs", "author": [ { "family_name": "Bodnarchuk", "given_name": "Maryna I.", "clpid": "Bodnarchuk-M-I" }, { "family_name": "Li", "given_name": "Liang", "clpid": "Li-Liang" }, { "family_name": "Fok", "given_name": "Alice", "clpid": "Fok-Alice" }, { "family_name": "Nachtergaele", "given_name": "Sigrid", "clpid": "Nachtergaele-S" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Talapin", "given_name": "Dmitri V.", "clpid": "Talapin-D-V" } ], "abstract": "We studied the self-assembly of inorganic nanocrystals (NCs) confined inside nanoliter droplets (plugs) into long-range ordered superlattices. We showed that a capillary microfluidic platform can be used for the optimization of growth conditions for NC superlattices and can provide insights into the kinetics of the NC assembly process. The utility of our approach was demonstrated by growing large (up to 200 \u03bcm) three-dimensional (3D) superlattices of various NCs, including Au, PbS, CdSe, and CoFe(2)O(4). We also showed that it is possible to grow 3D binary nanoparticle superlattices in the microfluidic plugs.", "doi": "10.1021/ja201129n", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2011-06-15", "series_number": "23", "volume": "133", "issue": "23", "pages": "8956-8960" }, { "id": "authors:caam1-ak276", "collection": "authors", "collection_id": "caam1-ak276", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160730642", "type": "article", "title": "Digital Isothermal Quantification of Nucleic Acids via Simultaneous Chemical Initiation of Recombinase Polymerase Amplification Reactions on SlipChip", "author": [ { "family_name": "Shen", "given_name": "Feng", "orcid": "0000-0002-4709-330X", "clpid": "Shen-Feng" }, { "family_name": "Davydova", "given_name": "Elena K.", "clpid": "Davydova-E-K" }, { "family_name": "Du", "given_name": "Wenbin", "orcid": "0000-0002-7401-1410", "clpid": "Du-Wenbin" }, { "family_name": "Kreutz", "given_name": "Jason E.", "clpid": "Kreutz-J-E" }, { "family_name": "Piepenburg", "given_name": "Olaf", "clpid": "Piepenburg-O" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "In this paper, digital quantitative detection of nucleic acids was achieved at the single-molecule level by chemical initiation of over one thousand sequence-specific, nanoliter isothermal amplification reactions in parallel. Digital polymerase chain reaction (digital PCR), a method used for quantification of nucleic acids, counts the presence or absence of amplification of individual molecules. However, it still requires temperature cycling, which is undesirable under resource-limited conditions. This makes isothermal methods for nucleic acid amplification, such as recombinase polymerase amplification (RPA), more attractive. A microfluidic digital RPA SlipChip is described here for simultaneous initiation of over one thousand nL-scale RPA reactions by adding a chemical initiator to each reaction compartment with a simple slipping step after instrument-free pipet loading. Two designs of the SlipChip, two-step slipping and one-step slipping, were validated using digital RPA. By using the digital RPA SlipChip, false-positive results from preinitiation of the RPA amplification reaction before incubation were eliminated. End point fluorescence readout was used for \"yes or no\" digital quantification. The performance of digital RPA in a SlipChip was validated by amplifying and counting single molecules of the target nucleic acid, methicillin-resistant Staphylococcus aureus (MRSA) genomic DNA. The digital RPA on SlipChip was also tolerant to fluctuations of the incubation temperature (37\u221242 \u00b0C), and its performance was comparable to digital PCR on the same SlipChip design. The digital RPA SlipChip provides a simple method to quantify nucleic acids without requiring thermal cycling or kinetic measurements, with potential applications in diagnostics and environmental monitoring under resource-limited settings. The ability to initiate thousands of chemical reactions in parallel on the nanoliter scale using solvent-resistant glass devices is likely to be useful for a broader range of applications.", "doi": "10.1021/ac200247e", "pmcid": "PMC3101872", "issn": "0003-2700", "publisher": "American Chemical Society", "publication": "Analytical Chemistry", "publication_date": "2011-05-01", "series_number": "9", "volume": "83", "issue": "9", "pages": "3533-3540" }, { "id": "authors:1dn94-4mv61", "collection": "authors", "collection_id": "1dn94-4mv61", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160722342", "type": "article", "title": "Complex Function by Design Using Spatially Pre-Structured Synthetic Microbial Communities: Degradation of Pentachlorophenol in the Presence of Hg(II)", "author": [ { "family_name": "Kim", "given_name": "Hyun Jung", "clpid": "Kim-Hyun-Jung" }, { "family_name": "Du", "given_name": "Wenbin", "orcid": "0000-0002-7401-1410", "clpid": "Du-Wenbin" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Naturally occurring microbes perform a variety of useful functions, with more complex processes requiring multiple functions performed by communities of multiple microbes. Synthetic biology via genetic engineering may be used to achieve desired multiple functions, e.g. multistep chemical and biological transformations, by adding genes to a single organism, but this is sometimes not possible due to incompatible metabolic requirements or not desirable in certain applications, especially in medical or environmental applications. Achieving multiple functions by mixing microbes that have not evolved to function together may not work due to competition of microbes, or lack of interactions among microbes. In nature, microbial communities are commonly spatially structured. Here, we tested whether spatial structure can be used to create a community of microbes that can perform a function they do not perform individually or when simply mixed. We constructed a core\u2013shell fiber with Sphingobium chlorophenolicum, a pentachlorophenol (PCP) degrader, in the core layer and Ralstonia metallidurans, a mercuric ion (Hg(II)) reducer, in the shell layer as a structured community using microfluidic laminar flow techniques. We applied a mixture of PCP and Hg(II) to either a simple well-mixed culture broth (i.e. the unstructured community) or the spatially structured core\u2013shell fibers. We found that without spatial structure, the community was unable to degrade PCP in the presence of Hg(II) because S. chlorophenolicum is sensitive to Hg(II). In contrast, with spatial structure in a core\u2013shell fiber system, S. chlorophenolicum in a core layer was protected by R. metallidurans deposited in a shell layer, and the community was able to completely remove both PCP and Hg(II) from a mixture. The appropriate size of the core\u2013shell fiber was determined by the Damk\u00f6hler number\u2014the timescale of removal of Hg(II) was on the same order of the timescale of diffusion of Hg(II) through the outer layer when the shell layer was on the order of B200 mm. Ultimately, with the ease of a child putting together 'Legos' to build a complex structure, using this approach one may be able to put together microorganisms to build communities that perform functions in vitro or even in vivo, e.g. as in a ''microbiome on a pill''.", "doi": "10.1039/C0IB00019A", "pmcid": "PMC3005148", "issn": "1757-9694", "publisher": "Royal Society of Chemistry", "publication": "Integrative Biology", "publication_date": "2011-02", "series_number": "2", "volume": "3", "issue": "2", "pages": "126-133" }, { "id": "authors:5s1fc-m2h24", "collection": "authors", "collection_id": "5s1fc-m2h24", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160729192", "type": "article", "title": "Microfluidics Using Spatially Defined Arrays of Droplets in One, Two, and Three Dimensions", "author": [ { "family_name": "Pompano", "given_name": "Rebecca R.", "clpid": "Pompano-R-R" }, { "family_name": "Liu", "given_name": "Weishan", "clpid": "Liu-Weishan" }, { "family_name": "Du", "given_name": "Wenbin", "orcid": "0000-0002-7401-1410", "clpid": "Du-Wenbin" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Spatially defined arrays of droplets differ from bulk emulsions in that droplets in arrays can be indexed on the basis of one or more spatial variables to enable identification, monitoring, and addressability of individual droplets. Spatial indexing is critical in experiments with hundreds to millions of unique compartmentalized microscale processes\u2014for example, in applications such as digital measurements of rare events in a large sample, high-throughput time-lapse studies of the contents of individual droplets, and controlled droplet-droplet interactions. This review describes approaches for spatially organizing and manipulating droplets in one-, two-, and three-dimensional structured arrays, including aspiration, laminar flow, droplet traps, the SlipChip, self-assembly, and optical or electrical fields. This review also presents techniques to analyze droplets in arrays and applications of spatially defined arrays, including time-lapse studies of chemical, enzymatic, and cellular processes, as well as further opportunities in chemical, biological, and engineering sciences, including perturbation/response experiments and personal and point-of-care diagnostics.", "doi": "10.1146/annurev.anchem.012809.102303", "issn": "1936-1327", "publisher": "Annual Reviews", "publication": "Annual Review of Analytical Chemistry", "publication_date": "2011", "volume": "4", "pages": "59-81" }, { "id": "authors:43mad-3yr47", "collection": "authors", "collection_id": "43mad-3yr47", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160730931", "type": "article", "title": "Digital PCR on a SlipChip", "author": [ { "family_name": "Shen", "given_name": "Feng", "orcid": "0000-0002-4709-330X", "clpid": "Shen-Feng" }, { "family_name": "Du", "given_name": "Wenbin", "orcid": "0000-0002-7401-1410", "clpid": "Du-Wenbin" }, { "family_name": "Kreutz", "given_name": "Jason E.", "clpid": "Kreutz-J-E" }, { "family_name": "Fok", "given_name": "Alice", "clpid": "Fok-Alice" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper describes a SlipChip to perform digital PCR in a very simple and inexpensive format. The fluidic path for introducing the sample combined with the PCR mixture was formed using elongated wells in the two plates of the SlipChip designed to overlap during sample loading. This fluidic path was broken up by simple slipping of the two plates that removed the overlap among wells and brought each well in contact with a reservoir preloaded with oil to generate 1280 reaction compartments (2.6 nL each) simultaneously. After thermal cycling, end-point fluorescence intensity was used to detect the presence of nucleic acid. Digital PCR on the SlipChip was tested quantitatively by using Staphylococcus aureus genomic DNA. As the concentration of the template DNA in the reaction mixture was diluted, the fraction of positive wells decreased as expected from the statistical analysis. No cross-contamination was observed during the experiments. At the extremes of the dynamic range of digital PCR the standard confidence interval determined using a normal approximation of the binomial distribution is not satisfactory. Therefore, statistical analysis based on the score method was used to establish these confidence intervals. The SlipChip provides a simple strategy to count nucleic acids by using PCR. It may find applications in research applications such as single cell analysis, prenatal diagnostics, and point-of-care diagnostics. SlipChip would become valuable for diagnostics, including applications in resource-limited areas after integration with isothermal nucleic acid amplification technologies and visual readout.", "doi": "10.1039/c004521g", "pmcid": "PMC2948063", "issn": "1473-0197", "publisher": "Royal Society of Chemistry", "publication": "Lab on a Chip", "publication_date": "2010-10-21", "series_number": "20", "volume": "10", "issue": "20", "pages": "2666-2672" }, { "id": "authors:05ksv-g7s05", "collection": "authors", "collection_id": "05ksv-g7s05", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160725940", "type": "article", "title": "Chemical Stimulation of the Arabidopsis thaliana Root using Multi-Laminar Flow on a Microfluidic Chip", "author": [ { "family_name": "Meier", "given_name": "Matthias", "orcid": "0000-0002-7179-4173", "clpid": "Meier-M-M-M" }, { "family_name": "Lucchetta", "given_name": "Elena M.", "clpid": "Lucchetta-E-M" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "In this article, we developed a \"plant on a chip\" microfluidic platform that can control the local chemical environment around live roots of Arabidopsis thaliana with high spatial resolution using multi-laminar flow. We characterized the flow profile around the Arabidopsis root, and verified that the shear forces within the device ([similar]10 dyne cm^\u22122) did not impede growth of the roots. Our platform was able to deliver stimuli to the root at a spatial resolution of 10\u2013800 \u00b5m. Further, the platform was validated by exposing desired regions of the root with a synthetic auxin derivative, 2,4-dichlorophenoxyacetic acid (2,4-D), and its inhibitor N-1-naphthylphthalamic acid (NPA). The response to the stimuli was observed using a DR5::GFP Arabidopsis line, where GFP expression is coupled to the auxin response regulator DR5. GFP expression in the root matched the position of the flow-focused stream containing 2,4-D. When the regions around the 2,4-D stimulus were exposed to the auxin transport inhibitor NPA, the active and passive transport mechanisms of auxin could be differentiated, as NPA blocks active cell-to-cell transport of auxin. Finally, we demonstrated that local 2,4-D stimulation in a [similar]10 \u00b5m root segment enhanced morphological changes such as epidermal hair growth. These experiments were proof-of-concept and agreed with the results expected based on known root biology, demonstrating that this \"root on a chip\" platform can be used to test how root development is affected by any chemical component of interest, including nitrogen, phosphate, salts, and other plant hormones.", "doi": "10.1039/c004629a", "pmcid": "PMC2912432", "issn": "1473-0197", "publisher": "Royal Society of Chemistry", "publication": "Lab on a Chip", "publication_date": "2010-08-21", "series_number": "16", "volume": "10", "issue": "16", "pages": "2147-2153" }, { "id": "authors:2shp1-1p995", "collection": "authors", "collection_id": "2shp1-1p995", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160723920", "type": "article", "title": "Dead-end filling of SlipChip evaluated theoretically and experimentally as a function of the surface chemistry and the gap size between the plates for lubricated and dry SlipChips", "author": [ { "family_name": "Li", "given_name": "Liang", "clpid": "Li-Liang" }, { "family_name": "Karymov", "given_name": "Mikhail A.", "clpid": "Karymov-M-A" }, { "family_name": "Nichols", "given_name": "Kevin P.", "clpid": "Nichols-K-P" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "In this paper, we describe a method to load a microfluidic device, the SlipChip, via dead-end filling. In dead-end filling, the lubricating fluid that fills the SlipChip after assembly is dissipated through the gap between the two plates of the SlipChip instead of flowing through an outlet at the end of the fluidic path. We describe a theoretical model and associated predictions of dead-end filling that takes into consideration the interfacial properties and the gap size between plates of SlipChips. In this method, filling is controlled by the balance of pressures: for filling to occur without leaking, the inlet pressure must be greater than the capillary pressure but less than the maximum sealing pressure. We evaluated our prediction with experiments, and our empirical results agreed well with theory. Internal reservoirs were designed to prevent evaporation during loading of multiple solutions. Solutions were first loaded one at a time into inlet reservoirs; by applying a single pressure source to the device, we were able to fill multiple fluidic paths simultaneously. We used this method to fill both lubricated and dry SlipChips. Dry-loaded SlipChips were fabricated from fluorinated ethylene propylene (FEP) by using hot embossing techniques, and were successfully filled and slipped to perform a simple chemical reaction. The SlipChip design was also modified to enable ease of filling by using multiple access holes to the inlet reservoir.", "doi": "10.1021/la101460z", "pmcid": "PMC2923639", "issn": "0743-7463", "publisher": "American Chemical Society", "publication": "Langmuir", "publication_date": "2010-07-20", "series_number": "14", "volume": "26", "issue": "14", "pages": "12465-12471" }, { "id": "authors:3m594-9ee16", "collection": "authors", "collection_id": "3m594-9ee16", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160719024", "type": "article", "title": "Molecular mechanism of MLL PHD3 and RNA recognition by the Cyp33 RRM domain", "author": [ { "family_name": "Hom", "given_name": "Robert A.", "clpid": "Hom-R-A" }, { "family_name": "Chang", "given_name": "Pei-Yun", "clpid": "Chang-Pei-Yun" }, { "family_name": "Roy", "given_name": "Siddhartha", "clpid": "Roy-Siddhartha" }, { "family_name": "Musselman", "given_name": "Catherine A.", "clpid": "Musselman-C-A" }, { "family_name": "Glass", "given_name": "Karen C.", "clpid": "Glass-K-C" }, { "family_name": "Selezneva", "given_name": "Anna I.", "clpid": "Selezneva-A-I" }, { "family_name": "Gozani", "given_name": "Or", "clpid": "Gozani-O" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Cleary", "given_name": "Michael L.", "clpid": "Cleary-M-L" }, { "family_name": "Kutateladze", "given_name": "Tatiana G.", "clpid": "Kutateladze-T-G" } ], "abstract": "The nuclear protein cyclophilin 33 (Cyp33) is a peptidyl-prolyl cis-trans isomerase that catalyzes cis-trans isomerization of the peptide bond preceding a proline and promotes folding and conformational changes in folded and unfolded proteins. The N-terminal RNA-recognition motif (RRM) domain of Cyp33 has been found to associate with the third plant homeodomain (PHD3) finger of the mixed lineage leukemia (MLL) proto-oncoprotein and a poly(A) RNA sequence. Here, we report a 1.9 A resolution crystal structure of the RRM domain of Cyp33 and describe the molecular mechanism of PHD3 and RNA recognition. The Cyp33 RRM domain folds into a five-stranded antiparallel beta-sheet and two alpha-helices. The RRM domain, but not the catalytic module of Cyp33, binds strongly to PHD3, exhibiting a 2 muM affinity as measured by isothermal titration calorimetry. NMR chemical shift perturbation (CSP) analysis and dynamics data reveal that the beta strands and the beta2-beta3 loop of the RRM domain are involved in the interaction with PHD3. Mutations in the PHD3-binding site or deletions in the beta2-beta3 loop lead to a significantly reduced affinity or abrogation of the interaction. The RNA-binding pocket of the Cyp33 RRM domain, mapped on the basis of NMR CSP and mutagenesis, partially overlaps with the PHD3-binding site, and RNA association is abolished in the presence of MLL PHD3. Full-length Cyp33 acts as a negative regulator of MLL-induced transcription and reduces the expression levels of MLL target genes MEIS1 and HOXA9. Together, these in vitro and in vivo data provide insight into the multiple functions of Cyp33 RRM and suggest a Cyp33-dependent mechanism for regulating the transcriptional activity of MLL.", "doi": "10.1016/j.jmb.2010.04.067", "pmcid": "PMC3204800", "issn": "0022-2836", "publisher": "Elsevier", "publication": "Journal of Molecular Biology", "publication_date": "2010-07-09", "series_number": "2", "volume": "400", "issue": "2", "pages": "145-154" }, { "id": "authors:4rsj5-qn162", "collection": "authors", "collection_id": "4rsj5-qn162", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160730786", "type": "article", "title": "Nanoliter Multiplex PCR Arrays on a SlipChip", "author": [ { "family_name": "Shen", "given_name": "Feng", "orcid": "0000-0002-4709-330X", "clpid": "Shen-Feng" }, { "family_name": "Du", "given_name": "Wenbin", "orcid": "0000-0002-7401-1410", "clpid": "Du-Wenbin" }, { "family_name": "Davydova", "given_name": "Elena K.", "clpid": "Davydova-E-K" }, { "family_name": "Karymov", "given_name": "Mikhail A.", "clpid": "Karymov-M-A" }, { "family_name": "Pandey", "given_name": "Janmajay", "clpid": "Pandey-J" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "The SlipChip platform was tested to perform highthroughput nanoliter multiplex PCR. The advantages of\nusing the SlipChip platform for multiplex PCR include the\nability to preload arrays of dry primers, instrument-free\nsample manipulation, small sample volume, and highthroughput capacity. The SlipChip was designed to preload one primer pair per reaction compartment and to\nscreen up to 384 different primer pairs with less than 30\nnanoliters of sample per reaction compartment. Both a\n40-well and a 384-well design of the SlipChip were tested\nfor multiplex PCR. In the geometries used here, the\nsample fluid was spontaneously compartmentalized into\ndiscrete volumes even before slipping of the two plates of\nthe SlipChip, but slipping introduced additional capabilities that made devices more robust and versatile. The\nwells of this SlipChip were designed to overcome potential\nproblems associated with thermal expansion. By using\ncircular wells filled with oil and overlapping them with\nsquare wells filled with the aqueous PCR mixture, a\ndroplet of aqueous PCR mixture was always surrounded\nby the lubricating fluid. In this design, during heating and\nthermal expansion, only oil was expelled from the compartment and leaking of the aqueous solution was prevented. Both 40-well and 384-well devices were found to\nbe free from cross-contamination, and end point fluorescence detection provided reliable readout. Multiple samples\ncould also be screened on the same SlipChip simultaneously. Multiplex PCR was validated on the 384-well\nSlipChip with 20 different primer pairs to identify 16\nbacterial and fungal species commonly presented in blood\ninfections. The SlipChip correctly identified five different\nbacterial or fungal species in separate experiments. In\naddition, the presence of the resistance gene mecA in\nmethicillin resistant Staphylococcus aureus (MRSA) was\nidentified. The SlipChip will be useful for applications\ninvolving PCR arrays and lays the foundation for new\nstrategies for diagnostics, point-of-care devices, and immobilization-based arrays.", "doi": "10.1021/ac1007249", "pmcid": "PMC2916686", "issn": "0003-2700", "publisher": "American Chemical Society", "publication": "Analytical Chemistry", "publication_date": "2010-06-01", "series_number": "11", "volume": "82", "issue": "11", "pages": "4606-4612" }, { "id": "authors:a1k69-5n905", "collection": "authors", "collection_id": "a1k69-5n905", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160723624", "type": "article", "title": "A Plug-Based Microfluidic System for Dispensing Lipidic Cubic Phase (LCP) Material Validated by Crystallizing Membrane Proteins in Lipidic Mesophases", "author": [ { "family_name": "Li", "given_name": "Liang", "clpid": "Li-Liang" }, { "family_name": "Fu", "given_name": "Qiang", "clpid": "Fu-Qiang" }, { "family_name": "Kors", "given_name": "Christopher A.", "clpid": "Kors-C-A" }, { "family_name": "Stewart", "given_name": "Lance", "clpid": "Stewart-L" }, { "family_name": "Nollert", "given_name": "Peter", "clpid": "Nollert-P" }, { "family_name": "Laible", "given_name": "Philip D.", "clpid": "Laible-P-D" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This article presents a plug-based microfluidic system to dispense nanoliter-volume plugs of lipidic cubic phase (LCP) material and subsequently merge the LCP plugs with aqueous plugs. This system was validated by crystallizing membrane proteins in lipidic mesophases, including LCP. This system allows for accurate dispensing of LCP material in nanoliter volumes, prevents inadvertent phase transitions that may occur due to dehydration by enclosing LCP in plugs, and is compatible with the traditional method of forming LCP material using a membrane protein sample, as shown by the successful crystallization of bacteriorhodopsin from Halobacterium salinarum. Conditions for the formation of LCP plugs were characterized and presented in a phase diagram. This system was also implemented using two different methods of introducing the membrane protein: (1) the traditional method of generating the LCP material using a membrane protein sample and (2) post LCP-formation incorporation (PLI), which involves making LCP material without protein, adding the membrane protein sample externally to the LCP material, and allowing the protein to diffuse into the LCP material or into other lipidic mesophases that may result from phase transitions. Crystals of bacterial photosynthetic reaction centers from Rhodobacter sphaeroides and Blastochloris viridis were obtained using PLI. The plug-based, LCP-assisted microfluidic system, combined with the PLI method for introducing membrane protein into LCP, should be useful for minimizing consumption of samples and broadening the screening of parameter space in membrane protein crystallization.", "doi": "10.1007/s10404-009-0512-8", "pmcid": "PMC2868346", "issn": "1613-4982", "publisher": "Springer", "publication": "Microfluidics and Nanofluidics", "publication_date": "2010-06", "series_number": "6", "volume": "8", "issue": "6", "pages": "789-798" }, { "id": "authors:966s2-k1w20", "collection": "authors", "collection_id": "966s2-k1w20", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160724379", "type": "article", "title": "SlipChip for immunoassays in nanoliter volumes", "author": [ { "family_name": "Liu", "given_name": "Weishan", "clpid": "Liu-Weishan" }, { "family_name": "Chen", "given_name": "Delai L.", "clpid": "Chen-Delai-L" }, { "family_name": "Du", "given_name": "Wenbin", "orcid": "0000-0002-7401-1410", "clpid": "Du-Wenbin" }, { "family_name": "Nichols", "given_name": "Kevin P.", "clpid": "Nichols-K-P" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This article describes a SlipChip-based approach to\nperform bead-based heterogeneous immunoassays with\nmultiple nanoliter-volume samples. As a potential device\nto analyze the output of the chemistrode, the performance\nof this platform was tested using low concentrations of\nbiomolecules. Two strategies to perform the immunoassay\nin the SlipChip were tested: (1) a unidirectional slipping\nmethod to combine the well containing a sample with a\nseries of wells preloaded with reagents and (2) a back-and-forth slipping method to introduce a series of reagents\nto a well containing the sample by reloading and slipping\nthe well containing the reagent. The SlipChips were\nfabricated with hydrophilic surfaces on the interior of the\nwells and with hydrophobic surfaces on the face of the\nSlipChip to enhance filling, transferring, and maintaining\naqueous solutions in shallow wells. Nanopatterning was\nused to increase the hydrophobic nature of the SlipChip\nsurface. Magnetic beads containing the capture antibody\nwere efficiently transferred between wells and washed by\nserial dilution. An insulin immunoenzymatic assay showed\na detection of limit of \u223c13 pM. A total of 48 droplets of\nnanoliter volume were analyzed in parallel, including an\non-chip calibration. The design of the SlipChip is flexible\nto accommodate other types of immunoassays, both\nheterogeneous and homogeneous. This work establishes\nthe possibility of using SlipChip-based immunoassays in\nsmall volumes for a range of possible applications, including analysis of plugs from a chemistrode, detection of\nmolecules from single cells, and diagnostic monitoring.", "doi": "10.1021/ac100044c", "pmcid": "PMC2885842", "issn": "0003-2700", "publisher": "American Chemical Society", "publication": "Analytical Chemistry", "publication_date": "2010-04-15", "series_number": "8", "volume": "82", "issue": "8", "pages": "3276-3282" }, { "id": "authors:nxcrk-vg436", "collection": "authors", "collection_id": "nxcrk-vg436", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160722961", "type": "article", "title": "Evolution of catalysts directed by genetic algorithms in a plug-based microfluidic device tested with oxidation of methane by oxygen", "author": [ { "family_name": "Kreutz", "given_name": "Jason E.", "clpid": "Kreutz-J-E" }, { "family_name": "Shukhaev", "given_name": "Anton", "clpid": "Shukhaev-A" }, { "family_name": "Du", "given_name": "Wenbin", "orcid": "0000-0002-7401-1410", "clpid": "Du-Wenbin" }, { "family_name": "Druskin", "given_name": "Sasha", "clpid": "Druskin-S" }, { "family_name": "Daugulis", "given_name": "Olafs", "clpid": "Daugulis-O" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper uses microfluidics to implement genetic algorithms (GA) to discover new homogeneous catalysts using the oxidation of methane by molecular oxygen as a model system. The parameters of the GA were the catalyst, a cocatalyst capable of using molecular oxygen as the terminal oxidant, and ligands that could tune the catalytic system. The GA required running hundreds of reactions to discover and optimize catalyst systems of high fitness, and microfluidics enabled these numerous reactions to be run in parallel. The small scale and volumes of microfluidics offer significant safety benefits. The microfluidic system included methods to form diverse arrays of plugs containing catalysts, introduce gaseous reagents at high pressure, run reactions in parallel, and detect catalyst activity using an in situ indicator system. Platinum(II) was identified as an active catalyst, and iron(II) and the polyoxometalate H5PMo10V2O40 (POM-V2) were identified as active cocatalysts. The Pt/Fe system was further optimized and characterized using NMR experiments. After optimization, turnover numbers of approximately 50 were achieved with approximately equal production of methanol and formic acid. The Pt/Fe system demonstrated the compatibility of iron with the entire catalytic cycle. This approach of GA-guided evolution has the potential to accelerate discovery in catalysis and other areas where exploration of chemical space is essential, including optimization of materials for hydrogen storage and CO2 capture and modifications.", "doi": "10.1021/ja909853x", "pmcid": "PMC2861856", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2010-03-10", "series_number": "9", "volume": "132", "issue": "9", "pages": "3128-3132" }, { "id": "authors:ybsrf-bq611", "collection": "authors", "collection_id": "ybsrf-bq611", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160734997", "type": "article", "title": "Microfluidic stochastic confinement enhances analysis of rare cells by isolating cells and creating high density environments for control of diffusible signals", "author": [ { "family_name": "Vincent", "given_name": "Meghan E.", "clpid": "Vincent-M-E" }, { "family_name": "Liu", "given_name": "Weishan", "clpid": "Liu-Weishan" }, { "family_name": "Haney", "given_name": "Elizabeth B.", "clpid": "Haney-E-B" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Rare cells can be difficult to analyze because they either occur in low numbers or coexist with a more abundant cell type, yet their detection is crucial for diagnosing disease and maintaining human health. In this tutorial review, we introduce the concept of microfluidic stochastic confinement for use in detection and analysis of rare cells. Stochastic confinement provides two advantages: (1) it separates rare single cells from the bulk mixture and (2) it allows signals to locally accumulate to a higher concentration around a single cell than in the bulk mixture. Microfluidics is an attractive method for implementing stochastic confinement because it provides simple handling of small volumes. We present technologies for microfluidic stochastic confinement that utilize both wells and droplets for the detection and analysis of single cells. We address how these microfluidic technologies have been used to observe new behavior, increase speed of detection, and enhance cultivation of rare cells. We discuss potential applications of microfluidic stochastic confinement to fields such as human diagnostics and environmental testing.", "doi": "10.1039/B917851A", "pmcid": "PMC2829723", "issn": "0306-0012", "publisher": "Royal Society of Chemistry", "publication": "Chemical Society Reviews", "publication_date": "2010-03", "series_number": "3", "volume": "39", "issue": "3", "pages": "974-984" }, { "id": "authors:x27g9-9tf08", "collection": "authors", "collection_id": "x27g9-9tf08", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160723429", "type": "article", "title": "User-Loaded SlipChip for Equipment-Free Multiplexed Nanoliter-Scale Experiments", "author": [ { "family_name": "Li", "given_name": "Liang", "clpid": "Li-Liang" }, { "family_name": "Du", "given_name": "Wenbin", "orcid": "0000-0002-7401-1410", "clpid": "Du-Wenbin" }, { "family_name": "Ismagilov", "given_name": "Rustem", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper describes a microfluidic approach to perform multiplexed nanoliter-scale experiments\nby combining a sample with multiple different reagents, each at multiple mixing ratios. This approach employs\na user-loaded, equipment-free SlipChip. The mixing ratios, characterized by diluting a fluorescent dye,\ncould be controlled by the volume of each of the combined wells. The SlipChip design was validated on an\n\u223c12 nL scale by screening the conditions for crystallization of glutaryl-CoA dehydrogenase from Burkholderia\npseudomallei against 48 different reagents; each reagent was tested at 11 different mixing ratios, for a\ntotal of 528 crystallization trials. The total consumption of the protein sample was \u223c10 \u00b5L. Conditions for\ncrystallization were successfully identified. The crystallization experiments were successfully scaled up in\nwell plates using the conditions identified in the SlipChip. Crystals were characterized by X-ray diffraction\nand provided a protein structure in a different space group and at a higher resolution than the structure\nobtained by conventional methods. In this work, this user-loaded SlipChip has been shown to reliably handle\nfluids of diverse physicochemical properties, such as viscosities and surface tensions. Quantitative\nmeasurements of fluorescent intensities and high-resolution imaging were straighforward to perform in\nthese glass SlipChips. Surface chemistry was controlled using fluorinated lubricating fluid, analogous to\nthe fluorinated carrier fluid used in plug-based crystallization. Thus, we expect this approach to be valuable\nin a number of areas beyond protein crystallization, especially those areas where droplet-based microfluidic\nsystems have demonstrated successes, including measurements of enzyme kinetics and blood coagulation,\ncell-based assays, and chemical reactions.", "doi": "10.1021/ja908555n", "pmcid": "PMC2802657", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2010-01-13", "series_number": "1", "volume": "132", "issue": "1", "pages": "106-111" }, { "id": "authors:crmb2-zjj74", "collection": "authors", "collection_id": "crmb2-zjj74", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160723281", "type": "article", "title": "Multiparameter Screening on SlipChip Used for Nanoliter Protein Crystallization Combining Free Interface Diffusion and Microbatch Methods", "author": [ { "family_name": "Li", "given_name": "Liang", "clpid": "Li-Liang" }, { "family_name": "Du", "given_name": "Wenbin", "orcid": "0000-0002-7401-1410", "clpid": "Du-Wenbin" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper describes two SlipChip-based approaches to protein crystallization: a SlipChip-based free interface diffusion (FID) method and a SlipChip-based composite method that simultaneously performs microbatch and FID crystallization methods in a single device. The FID SlipChip was designed to screen multiple reagents, each at multiple diffusion equilibration times, and was validated by screening conditions for crystallization of two proteins, enoyl-CoA hydratase from Mycobacterium tuberculosis and dihydrofolate reductase/thymidylate synthase from Babesia bovis, against 48 different reagents at five different equilibration times each, consuming 12 \u00b5L of each protein for a total of 480 experiments using three SlipChips. The composite SlipChip was designed to screen multiple reagents, each at multiple mixing ratios and multiple equilibration times, and was validated by screening conditions for crystallization of two proteins, enoylCoA hydratase from Mycobacterium tuberculosis and dihydrofolate reductase/thymidylate synthase from Babesia bovis. To prevent cross-contamination while keeping the solution in the neck channels for FID stable, the plates of the SlipChip were etched with a pattern of nanowells. This nanopattern was used to increase the contact angle of aqueous solutions on the surface of the silanized glass. The composite SlipChip increased the number of successful crystallization conditions and identified more conditions for crystallization than separate FID and microbatch screenings. Crystallization experiments were scaled up in well plates using conditions identified during the SlipChip screenings, and X-ray diffraction data were obtained to yield the protein structure of dihydrofolate reductase/thymidylate synthase at 1.95 \u00c5 resolution. This free-interface diffusion approach provides a convenient and high-throughput method of setting up gradients in microfluidic devices and may find additional applications in cell-based assays.", "doi": "10.1021/ja908558m", "pmcid": "PMC2805062", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2010-01-13", "series_number": "1", "volume": "132", "issue": "1", "pages": "112-119" }, { "id": "authors:daxyw-2tt16", "collection": "authors", "collection_id": "daxyw-2tt16", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160723789", "type": "article", "title": "Protein Crystallization using Microfluidic Technologies Based on Valves, Droplets, and SlipChip", "author": [ { "family_name": "Li", "given_name": "Liang", "clpid": "Li-Liang" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "To obtain protein crystals, researchers must search for conditions in multidimensional chemical space. Empirically, thousands of crystallization experiments are carried out to screen various precipitants at multiple concentrations. Microfluidics can manipulate fluids on a nanoliter scale, and it affects crystallization twofold. First, it miniaturizes the experiments that can currently be done on a larger scale and enables crystallization of proteins that are available only in small amounts. Second, it offers unique experimental approaches that are difficult or impossible to implement on a larger scale. Ongoing development of microfluidic techniques and their integration with protein production, characterization, and in situ diffraction promises to accelerate the progress of structural biology.", "doi": "10.1146/annurev.biophys.050708.133630", "issn": "1936-122X", "publisher": "Annual Reviews", "publication": "Annual Review of Biophysics", "publication_date": "2010", "volume": "39", "pages": "139-158" }, { "id": "authors:n51hm-56854", "collection": "authors", "collection_id": "n51hm-56854", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160724842", "type": "article", "title": "The Endo-siRNA Pathway Is Essential for Robust Development of the Drosophila Embryo", "author": [ { "family_name": "Lucchetta", "given_name": "Elena M.", "clpid": "Lucchetta-E-M" }, { "family_name": "Carthew", "given_name": "Richard W.", "clpid": "Carthew-R-W" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Background: Robustness to natural temperature fluctuations is critical to proper development in embryos and to cellular\nfunctions in adult organisms. However, mechanisms and pathways which govern temperature compensation remain largely\nunknown beyond circadian rhythms. Pathways which ensure robustness against temperature fluctuations may appear to be\nnonessential under favorable, uniform environmental conditions used in conventional laboratory experiments where there\nis little variation for which to compensate. The endo-siRNA pathway, which produces small double-stranded RNAs in\nDrosophila, appears to be nonessential for robust development of the embryo under ambient uniform temperature and to\nbe necessary only for viral defense. Embryos lacking a functional endo-siRNA pathway develop into phenotypically normal\nadults. However, we hypothesized that small RNAs may regulate the embryo's response to temperature, as a\nribonucleoprotein complex has been previously shown to mediate mammalian cell response to heat shock.\nPrincipal Findings: Here, we show that the genes DICER-2 and ARGONAUTE2, which code for integral protein components\nof the endo-siRNA pathway, are essential for robust development and temperature compensation in the Drosophila embryo\nwhen exposed to temperature perturbations. The regulatory functions of DICER-2 and ARGONAUTE2 were uncovered by\nusing microfluidics to expose developing Drosophila embryos to a temperature step, in which each half of the embryo\ndevelops at a different temperature through developmental cycle 14. Under this temperature perturbation, dicer-2 or\nargonaute2 embryos displayed abnormal segmentation. The abnormalities in segmentation are presumably due to the\ninability of the embryo to compensate for temperature-induced differences in rate of development and to coordinate\ndevelopmental timing in the anterior and posterior halves. A deregulation of the length of nuclear division cycles 10\u201314 is\nalso observed in dicer-2 embryos at high temperatures.\nConclusions: Results presented herein uncover a novel function of the endo-siRNA pathway in temperature compensation\nand cell cycle regulation, and we hypothesize that the endo-siRNA pathway may regulate the degradation of maternal cell\ncycle regulators. Endo-siRNAs may have a more general role buffering against environmental perturbations in other\norganisms.", "doi": "10.1371/journal.pone.0007576", "pmcid": "PMC2761733", "issn": "1932-6203", "publisher": "Public Library of Science", "publication": "PLoS ONE", "publication_date": "2009-10-23", "series_number": "10", "volume": "4", "issue": "10", "pages": "e7576" }, { "id": "authors:5j8at-mxj77", "collection": "authors", "collection_id": "5j8at-mxj77", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160731264", "type": "article", "title": "Confinement Regulates Complex Biochemical Networks: Initiation of Blood Clotting by \"Diffusion Acting\"", "author": [ { "family_name": "Shen", "given_name": "Feng", "orcid": "0000-0002-4709-330X", "clpid": "Shen-Feng" }, { "family_name": "Pompano", "given_name": "Rebecca R.", "clpid": "Pompano-Rebecca-R" }, { "family_name": "Kastrup", "given_name": "Christian J.", "clpid": "Kastrup-Christian-J" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This study shows that environmental confinement strongly affects the activation of nonlinear reaction networks, such as blood coagulation (clotting), by small quantities of activators. Blood coagulation is sensitive to the local concentration of soluble activators, initiating only when the activators surpass a threshold concentration, and therefore is regulated by mass transport phenomena such as flow and diffusion. Here, diffusion was limited by decreasing the size of microfluidic chambers, and it was found that microparticles carrying either the classical stimulus, tissue factor, or a bacterial stimulus, Bacillus cereus, initiated coagulation of human platelet-poor plasma only when confined. A simple analytical argument and numerical model were used to describe the mechanism for this phenomenon: confinement causes diffusible activators to accumulate locally and surpass the threshold concentration. To interpret the results, a dimensionless confinement number, Cn, was used to describe whether a stimulus was confined, and a Damkohler number, Da_(2), was used to describe whether a subthreshold stimulus could initiate coagulation. In the context of initiation of coagulation by bacteria, this mechanism can be thought of as \"diffusion acting\", which is distinct from \"diffusion sensing\". The ability of confinement and diffusion acting to change the outcome of coagulation suggests that confinement should also regulate other biological \"on\" and \"off\" processes that are controlled by thresholds.", "doi": "10.1016/j.bpj.2009.08.004", "pmcid": "PMC2764071", "issn": "0006-3495", "publisher": "Biophysical Society", "publication": "Biophysical Journal", "publication_date": "2009-10-21", "series_number": "8", "volume": "97", "issue": "8", "pages": "2137-2145" }, { "id": "authors:9xtqf-e0m16", "collection": "authors", "collection_id": "9xtqf-e0m16", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160729461", "type": "article", "title": "Structural and spectropotentiometric analysis of Blastochloris viridis heterodimer mutant reaction center", "author": [ { "family_name": "Ponomarenko", "given_name": "Nina S.", "clpid": "Ponomarenko-Nina-S" }, { "family_name": "Li", "given_name": "Liang", "clpid": "Li-Liang" }, { "family_name": "Marino", "given_name": "Antony R.", "clpid": "Marino-Antony-R" }, { "family_name": "Tereshko", "given_name": "Valentina", "clpid": "Tereshko-Valentina" }, { "family_name": "Ostafin", "given_name": "Agnes", "clpid": "Ostafin-Agnes" }, { "family_name": "Popova", "given_name": "Julia A.", "clpid": "Popova-Julia-A" }, { "family_name": "Bylina", "given_name": "Edward J.", "clpid": "Bylina-Edward-J" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Norris", "given_name": "James R., Jr.", "clpid": "Norris-James-R-Jr" } ], "abstract": "Heterodimer mutant reaction centers (RCs) of Blastochloris viridis were crystallized using microfluidic\ntechnology. In this mutant, a leucine residue replaced the histidine residue which had acted as a fifth ligand\nto the bacteriochlorophyll (BChl) of the primary electron donor dimer M site (HisM200). With the loss of the\nhistidine-coordinated Mg, one bacteriochlorophyll of the special pair was converted into a bacteriopheophytin (BPhe), and the primary donor became a heterodimer supermolecule. The crystals had dimensions\n400 \u00d7 100 \u00d7100 \u03bcm, belonged to space group P43212, and were isomorphous to the ones reported earlier for\nthe wild type (WT) strain. The structure was solved to a 2.5 \u00c5 resolution limit. Electron-density maps\nconfirmed the replacement of the histidine residue and the absence of Mg. Structural changes in the\nheterodimer mutant RC relative to the WT included the absence of the water molecule that is typically\npositioned between the M side of the primary donor and the accessory BChl, a slight shift in the position of\namino acids surrounding the site of the mutation, and the rotation of the M194 phenylalanine. The\ncytochrome subunit was anchored similarly as in the WT and had no detectable changes in its overall\nposition. The highly conserved tyrosine L162, located between the primary donor and the highest potential\nheme C380, revealed only a minor deviation of its hydroxyl group. Concomitantly to modification of the BChl\nmolecule, the redox potential of the heterodimer primary donor increased relative to that of the WT\norganism (772 mV vs. 517 mV). The availability of this heterodimer mutant and its crystal structure provides\nopportunities for investigating changes in light-induced electron transfer that reflect differences in redox\ncascades.", "doi": "10.1016/j.bbamem.2009.06.006", "pmcid": "PMC2752317", "issn": "0005-2736", "publisher": "Elsevier", "publication": "Biochimica et Biophysica Acta - Biomembranes", "publication_date": "2009-09", "volume": "1788", "pages": "1822-1831" }, { "id": "authors:488kj-c6409", "collection": "authors", "collection_id": "488kj-c6409", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160718040", "type": "article", "title": "SlipChip", "author": [ { "family_name": "Du", "given_name": "Wenbin", "orcid": "0000-0002-7401-1410", "clpid": "Du-Wenbin" }, { "family_name": "Li", "given_name": "Liang", "clpid": "Li-Liang" }, { "family_name": "Nichols", "given_name": "Kevin P.", "clpid": "Nichols-Kevin-P" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "The SlipChip is a microfluidic device designed to perform multiplexed microfluidic reactions without\npumps or valves. The device has two plates in close contact. The bottom plate contains wells preloaded\nwith many reagents; in this paper plates with 48 reagents were used. These wells are covered by the top\nplate that acts as a lid for the wells with reagents. The device also has a fluidic path, composed of ducts\nin the bottom plate and wells in the top plate, which is connected only when the top and bottom plate\nare aligned in a specific configuration. Sample can be added into the fluidic path, filling both wells and\nducts. Then, the top plate is ''slipped'', or moved, relative to the bottom plate so the complementary\npatterns of wells in both plates overlap, exposing the sample-containing wells of the top plate to the\nreagent-containing wells of the bottom plate, and enabling diffusion and reactions. Between the two\nplates, a lubricating layer of fluorocarbon was used to facilitate relative motion of the plates. This paper\nimplements this approach on a nanoliter scale using devices fabricated in glass. Stability of preloaded\nsolutions, control of loading, and lack of cross-contamination were tested using fluorescent dyes.\nFunctionality of the device was illustrated via crystallization of a model membrane protein. Fabrication\nof this device is simple and does not require a bonding step. This device requires no pumps or valves and\nis applicable to resource-poor settings. Overall, this device should be valuable for multiplexed\napplications that require exposing one sample to many reagents in small volumes. One may think of the\nSlipChip as an easy-to-use analogue of a preloaded multi-well plate, or a preloaded liquid-phase\nmicroarray.", "doi": "10.1039/B908978K", "pmcid": "PMC2719824", "issn": "1473-0197", "publisher": "Royal Society of Chemistry", "publication": "Lab on a Chip", "publication_date": "2009-08-21", "series_number": "16", "volume": "9", "issue": "16", "pages": "2286-2292" }, { "id": "authors:xz0jq-jkp15", "collection": "authors", "collection_id": "xz0jq-jkp15", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160724539", "type": "article", "title": "Isolation, incubation, and parallel functional testing and identification by FISH of rare microbial single-copy cells from multi-species mixtures using the combination of chemistrode and stochastic confinement", "author": [ { "family_name": "Liu", "given_name": "Weishan", "clpid": "Liu-Weishan" }, { "family_name": "Kim", "given_name": "Hyun Jung", "clpid": "Kim-Hyun-Jung" }, { "family_name": "Lucchetta", "given_name": "Elena M.", "clpid": "Lucchetta-Elena-M" }, { "family_name": "Du", "given_name": "Wenbin", "orcid": "0000-0002-7401-1410", "clpid": "Du-Wenbin" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper illustrates a plug-based microfluidic approach combining the technique of the chemistrode and the principle of stochastic confinement, which can be used to i) starting from a mixture of cells, stochastically isolate single cells into plugs, ii) incubate the plugs to grow clones of the individual cells without competition among different clones, iii) split the plugs into arrays of identical daughter plugs, where each plug contained clones of the original cell, and iv) analyze each array by an independent technique, including cellulase assays, cultivation, cryo-preservation, Gram staining, and Fluorescence In Situ Hybridization (FISH). Functionally, this approach is equivalent to simultaneously assaying the clonal daughter cells by multiple killing and non-killing methods. A new protocol for single-cell FISH, a killing method, was developed to identify isolated cells of Paenibacillus curdlanolyticus in one array of daughter plugs using a 16S rRNA probe, Pc196. At the same time, live copies of P. curdlanolyticus in another array were obtained for cultivation. Among technical advances, this paper reports a chemistrode that enables sampling of nanoliter volumes directly from environmental specimens, such as soil slurries. In addition, a method for analyzing plugs is described: an array of droplets is deposited on the surface, and individual plugs are injected into the droplets of the surface array to induce a reaction and enable microscopy without distortions associated with curvature of plugs. The overall approach is attractive for identifying rare, slow growing microorganisms and would complement current methods to cultivate unculturable microbes from environmental samples.", "doi": "10.1039/b904958d", "pmcid": "PMC2719823", "issn": "1473-0197", "publisher": "Royal Society of Chemistry", "publication": "Lab on a Chip", "publication_date": "2009-08-07", "series_number": "15", "volume": "9", "issue": "15", "pages": "2153-2162" }, { "id": "authors:89xab-bww64", "collection": "authors", "collection_id": "89xab-bww64", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160716895", "type": "article", "title": "Using TIRF microscopy to quantify and confirm efficient mass transfer at the substrate surface of the chemistrode", "author": [ { "family_name": "Chen", "given_name": "Delai", "clpid": "Chen-Delai-L" }, { "family_name": "Du", "given_name": "Wenbin", "orcid": "0000-0002-7401-1410", "clpid": "Du-Wenbin" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper describes experiments for characterizing mass transfer\nat the hydrophilic surface of the substrate in a chemistrode. The chemistrode\nuses microfluidic plugs to deliver pulses of chemicals to a substrate with\nhigh temporal resolution, which requires efficient mass transfer between the\nwetting layer and the hydrophilic surface of the substrate. Here, total internal\nreflection fluorescence microscopy (TIRFM) was used to image the hydrophilic\nsurface of the substrate as plugs were made to flow over it. The surface of\nthe substrate was rapidly saturated with a fluorescent dye as the fluroesecent\nplugs passed over the substrate, confirming effective mass transfer between the\nwetting layer and the surface of the substrate. The dynamics of saturation are\nconsistent from cycle to cycle, indicating that the chemistrode can stimulate\nsurfaces with high reproducibility. The number of plugs required to reach\n90% saturation of the hydrophilic surface of the substrate, \u03c6(90%), only\nweakly depended on experimental conditions (the P\u00e9clet number or the capillary\nnumber). Furthermore, over a wide range of operating conditions, \u03c6(90%) was\nless than 4. These results are useful for improving the chemistrode and for\nunderstanding other phenomena that involve diffusional transfer in multiphase\nor recirculating flows near surfaces.", "doi": "10.1088/1367-2630/11/7/075017", "pmcid": "PMC2757094", "issn": "1367-2630", "publisher": "IOP", "publication": "New Journal of Physics", "publication_date": "2009-07", "series_number": "7", "volume": "11", "issue": "7", "pages": "Art. No. 075017" }, { "id": "authors:m8f63-s1c02", "collection": "authors", "collection_id": "m8f63-s1c02", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160722626", "type": "article", "title": "Laterally Mobile, Functionalized Self-Assembled Monolayers at the Fluorous-Aqueous Interface in a Plug-Based Microfluidic System: Characterization and Testing with Membrane Protein Crystallization", "author": [ { "family_name": "Kreutz", "given_name": "Jason E.", "clpid": "Kreutz-Jason-E" }, { "family_name": "Li", "given_name": "Liang", "clpid": "Li-Liang" }, { "family_name": "Roach", "given_name": "L. Spencer", "clpid": "Roach-L-Spencer" }, { "family_name": "Hatakeyama", "given_name": "Takuji", "clpid": "Hatakeyama-Takuji" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper describes a method to generate functionalizable, mobile self-assembled monolayers (SAMs) in plug-based microfluidics. Control of interfaces is advancing studies of biological interfaces, heterogeneous reactions, and nanotechnology. SAMs have been useful for such studies, but they are not laterally mobile. Lipid-based methods, though mobile, are not easily amenable to setting up the hundreds of experiments necessary for crystallization screening. Here we demonstrate a method, complementary to current SAM and lipid methods, for rapidly generating mobile, functionalized SAMs. This method relies on plugs, droplets surrounded by a fluorous carrier fluid, to rapidly explore chemical space. Specifically, we implemented his-tag binding chemistry to design a new fluorinated amphiphile, RfNTA, using an improved one-step synthesis of RfOEG under Mitsunobu conditions. RfNTA introduces specific binding of protein at the fluorous\u2212aqueous interface, which concentrates and orients proteins at the interface, even in the presence of other surfactants. We then applied this approach to the crystallization of a his-tagged membrane protein, Reaction Center from Rhodobacter sphaeroides, performed 2400 crystallization trials, and showed that this approach can increase the range of crystal-producing conditions, the success rate at a given condition, the rate of nucleation, and the quality of the crystal formed.", "doi": "10.1021/ja808697e", "pmcid": "PMC2741014", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2009-05-06", "series_number": "17", "volume": "131", "issue": "17", "pages": "6042-6043" }, { "id": "authors:2nvy5-vw809", "collection": "authors", "collection_id": "2nvy5-vw809", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160724691", "type": "article", "title": "Dynamics of Coalescence of Plugs with a Hydrophilic Wetting Layer Induced by Flow in a Microfluidic Chemistrode", "author": [ { "family_name": "Liu", "given_name": "Ying", "clpid": "Liu-Ying" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This manuscript analyzes the dynamics of coalescence of an incoming aqueous plug with a wetting layer above a hydrophilic surface in the chemistrode. The chemistrode is a recently described (Chen, D.; Du, W.; Liu, Y.; Liu, W.; Kuznetsov, A.; Mendez, F. E.; Philipson, L. H.; Ismagilov, R. F. Proc. Natl. Acad. Sci. U.S.A. 2008, 105, 16843-16848) microfluidic analogue of an electrode, but operating at the chemical rather than electrical level, developed with the aim of capturing local stimulus-response processes in chemistry and biology. The chemistrode consists of open-ended V-shaped microfluidic channels that can be brought into contact with a chemical or biological hydrophilic substrate. The chemistrode relies on multiphase aqueous/fluorous flow and uses plugs to achieve high temporal resolution of stimulation and sampling. Coalescence of the incoming plugs, containing the stimuli, with the liquid in the wetting layer is required for chemical exchange to take place in the chemistrode. Here, we investigate the system with triethyleneglycol mono1H, 1H-perfluorooctyl ether RfOEG as the surfactant. This surfactant was necessary to prevent nonspecific absorption of proteins to the aqueous fluorous interface and to ensure biocompatibility of the system, but too much surfactant increased the barrier for coalescence. In this system, coalescence was controlled by the capillary number. At a higher value of the capillary number, coalescence took more time, and deformation of the interface of the incoming plug and the wetting layer was more significant. Above a critical capillary number, coalescence did not occur between the incoming plug and the wetting layer. The critical capillary number was an increasing function of surface tension but was independent of viscosity ratio. Coalescence was surprisingly reproducible, presumably because film rupture during coalescence was reliably initiated at the hydrophilic substrate. These results are useful in rational operation of the chemistrode and also provide an experimental description of deformation, film drainage, and coalescence of surfactant-coated droplets in an external flow field.", "doi": "10.1021/la803518b", "issn": "0743-7463", "publisher": "American Chemical Society", "publication": "Langmuir", "publication_date": "2009-03-03", "series_number": "5", "volume": "25", "issue": "5", "pages": "2854-2859" }, { "id": "authors:rhnsn-z3k82", "collection": "authors", "collection_id": "rhnsn-z3k82", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160725760", "type": "article", "title": "Plug-Based Microfluidics with Defined Surface Chemistry to Miniaturize and Control Aggregation of Amyloidogenic Peptides", "author": [ { "family_name": "Meier", "given_name": "Matthias", "orcid": "0000-0002-7179-4173", "clpid": "Meier-Matthias-M-M" }, { "family_name": "Kennedy-Darling", "given_name": "Julia", "clpid": "Kennedy-Darling-J" }, { "family_name": "Choi", "given_name": "Se Hoon", "clpid": "Choi-Se-Hoon" }, { "family_name": "Norstrom", "given_name": "Eric M.", "clpid": "Norstrom-Eric-M" }, { "family_name": "Sisodia", "given_name": "Sangram S.", "clpid": "Sisodia-Sangram-S" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Small with control: For miniaturization of protein aggregation experiments the interfacial chemistry must be controlled to avoid protein aggregation caused by interfacial adsorption. Plug-based microfluidics with defined surface chemistry (see schematic picture) can then be used to perform hundreds of aggregation experiments with volume-limited samples, such as cerebrospinal fluid from mice.", "doi": "10.1002/anie.200805225", "pmcid": "PMC2675571", "issn": "1433-7851", "publisher": "Wiley", "publication": "Angewandte Chemie International Edition", "publication_date": "2009-02-09", "series_number": "8", "volume": "48", "issue": "8", "pages": "1487-1489" }, { "id": "authors:6c334-gzw38", "collection": "authors", "collection_id": "6c334-gzw38", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160716355", "type": "article", "title": "Microfluidic confinement of single cells of bacteria in small volumes initiates high density behavior of quorum sensing and growth and reveals its variability", "author": [ { "family_name": "Boedicker", "given_name": "James Q.", "orcid": "0000-0003-4107-3719", "clpid": "Boedicker-James-Q" }, { "family_name": "Vincent", "given_name": "Meghan E.", "clpid": "Vincent-Meghan-E" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "One is a quorum: As few as one to three cells of Pseudomonas aeruginosa bacteria are confined in small volumes by the use of microfluidics. These small numbers of cells are able to activate quorum sensing (QS) pathways and achieve QS-dependent growth. The results also show that at low numbers of cells, initiation of QS is highly variable within a clonal population.", "doi": "10.1002/anie.200901550", "pmcid": "PMC2748941", "issn": "1433-7851", "publisher": "Wiley", "publication": "Angewandte Chemie International Edition", "publication_date": "2009", "series_number": "32", "volume": "48", "issue": "32", "pages": "5908-5911" }, { "id": "authors:c8zsy-h1051", "collection": "authors", "collection_id": "c8zsy-h1051", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160720712", "type": "article", "title": "Spatial localization of bacteria controls coagulation of human blood by 'quorum acting'", "author": [ { "family_name": "Kastrup", "given_name": "Christian J.", "clpid": "Kastrup-Christian-J" }, { "family_name": "Boedicker", "given_name": "James Q.", "orcid": "0000-0003-4107-3719", "clpid": "Boedicker-James-Q" }, { "family_name": "Pomerantsev", "given_name": "Andrei P.", "clpid": "Pomerantsev-Andrei-P" }, { "family_name": "Moayeri", "given_name": "Mahtab", "clpid": "Moayeri-Mahtab" }, { "family_name": "Bian", "given_name": "Yao", "clpid": "Bian-Yao" }, { "family_name": "Pompano", "given_name": "Rebecca R.", "clpid": "Pompano-Rebecca-R" }, { "family_name": "Kline", "given_name": "Timothy R.", "clpid": "Kline-Timothy-R" }, { "family_name": "Sylvestre", "given_name": "Patricia", "clpid": "Sylvestre-Patricia" }, { "family_name": "Shen", "given_name": "Feng", "orcid": "0000-0002-4709-330X", "clpid": "Shen-Feng" }, { "family_name": "Leppla", "given_name": "Stephen H.", "clpid": "Leppia-Stephen-H" }, { "family_name": "Tang", "given_name": "Wei-Jen", "clpid": "Tang-Wei-Jen" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Blood coagulation often accompanies bacterial infections and sepsis and is generally accepted as a consequence of immune\nresponses. Though many bacterial species can directly activate individual coagulation factors, they have not been shown to\ndirectly initiate the coagulation cascade that precedes clot formation. Here we demonstrated, using microfluidics and surface\npatterning, that the spatial localization of bacteria substantially affects coagulation of human and mouse blood and plasma.\nBacillus cereus and Bacillus anthracis, the anthrax-causing pathogen, directly initiated coagulation of blood in minutes when\nbacterial cells were clustered. Coagulation of human blood by B. anthracis required secreted zinc metalloprotease InhA1, which\nactivated prothrombin and factor X directly (not via factor XII or tissue factor pathways). We refer to this mechanism as 'quorum\nacting' to distinguish it from quorum sensing\u2014it does not require a change in gene expression, it can be rapid and it can be\nindependent of bacterium-to-bacterium communication.", "doi": "10.1038/nchembio.124", "pmcid": "PMC2651025", "issn": "1552-4450", "publisher": "Nature Publishing Group", "publication": "Nature Chemical Biology", "publication_date": "2008-12", "series_number": "12", "volume": "4", "issue": "12", "pages": "742-750" }, { "id": "authors:aps6x-9gh14", "collection": "authors", "collection_id": "aps6x-9gh14", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160722189", "type": "article", "title": "Defined spatial structure stabilizes a synthetic multispecies bacterial community", "author": [ { "family_name": "Kim", "given_name": "Hyun Jung", "clpid": "Kim-Hyun-Jung" }, { "family_name": "Boedicker", "given_name": "James Q.", "orcid": "0000-0003-4107-3719", "clpid": "Boedicker-J-Q" }, { "family_name": "Choi", "given_name": "Jang Wook", "orcid": "0000-0001-8783-0901", "clpid": "Choi-Jang-Wook" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper shows that for microbial communities, ''fences make\ngood neighbors.'' Communities of soil microorganisms perform\ncritical functions: controlling climate, enhancing crop production,\nand remediation of environmental contamination. Microbial communities in the oral cavity and the gut are of high biomedical\ninterest. Understanding and harnessing the function of these\ncommunities is difficult: artificial microbial communities in the\nlaboratory become unstable because of ''winner-takes-all'' competition among species. We constructed a community of three\ndifferent species of wild-type soil bacteria with syntrophic interactions using a microfluidic device to control spatial structure and\nchemical communication. We found that defined microscale spatial\nstructure is both necessary and sufficient for the stable coexistence\nof interacting bacterial species in the synthetic community. A\nmathematical model describes how spatial structure can balance\nthe competition and positive interactions within the community,\neven when the rates of production and consumption of nutrients\nby species are mismatched, by exploiting nonlinearities of these\nprocesses. These findings provide experimental and modeling\nevidence for a class of communities that require microscale spatial\nstructure for stability, and these results predict that controlling\nspatial structure may enable harnessing the function of natural and\nsynthetic multispecies communities in the laboratory.", "doi": "10.1073/pnas.0807935105", "pmcid": "PMC2587551", "issn": "0027-8424", "publisher": "National Academy of Sciences", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "publication_date": "2008-11-25", "series_number": "47", "volume": "105", "issue": "47", "pages": "18188-18193" }, { "id": "authors:c89dv-f6h57", "collection": "authors", "collection_id": "c89dv-f6h57", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160725343", "type": "article", "title": "A Precise Bicoid Gradient Is Nonessential during Cycles 11\u201313 for Precise Patterning in the Drosophila Blastoderm", "author": [ { "family_name": "Lucchetta", "given_name": "Elena M.", "clpid": "Lucchetta-E-M" }, { "family_name": "Vincent", "given_name": "Meghan E.", "clpid": "Vincent-M-E" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Background: During development, embryos decode maternal morphogen inputs into highly precise zygotic gene\nexpression. The discovery of the morphogen Bicoid and its profound effect on developmental programming in the\nDrosophila embryo has been a cornerstone in understanding the decoding of maternal inputs. Bicoid has been described as\na classical morphogen that forms a concentration gradient along the antero-posterior axis of the embryo by diffusion and\ninitiates expression of target genes in a concentration-dependent manner in the syncytial blastoderm. Recent work has\nemphasized the stability of the Bicoid gradient as a function of egg length and the role of nuclear dynamics in maintaining\nthe Bicoid gradient. Bicoid and nuclear dynamics were observed but not modulated under the ideal conditions used\npreviously. Therefore, it has not been tested explicitly whether a temporally stable Bicoid gradient prior to cellularization is\nrequired for precise patterning.\nPrincipal Findings: Here, we modulate both nuclear dynamics and the Bicoid gradient using laminar flows of different\ntemperature in a microfluidic device to determine if stability of the Bicoid gradient prior to cellularization is essential for\nprecise patterning. Dramatic motion of both cytoplasm and nuclei was observed prior to cellularization, and the Bicoid\ngradient was disrupted by nuclear motion and was highly abnormal as a function of egg length. Despite an abnormal Bicoid\ngradient during cycles 11\u201313, Even-skipped patterning in these embryos remained precise.\nConclusions: These results indicate that the stability of the Bicoid gradient as a function of egg length is nonessential\nduring syncytial blastoderm stages. Further, presumably no gradient formed by simple diffusion on the scale of egg length\ncould be responsible for the robust antero-posterior patterning observed, as severe cytoplasmic and nuclear motion would\ndisrupt such a gradient. Additional mechanisms for how the embryo could sense its dimensions and interpret the Bicoid\ngradient are discussed.", "doi": "10.1371/journal.pone.0003651", "pmcid": "PMC2578877", "issn": "1932-6203", "publisher": "Public Library of Science", "publication": "PLoS ONE", "publication_date": "2008-11-07", "series_number": "11", "volume": "3", "issue": "11", "pages": "e3651" }, { "id": "authors:5mws5-zjr82", "collection": "authors", "collection_id": "5mws5-zjr82", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160717046", "type": "article", "title": "The chemistrode: A droplet-based microfluidic device for stimulation and recording with high temporal, spatial, and chemical resolution", "author": [ { "family_name": "Chen", "given_name": "Delai", "clpid": "Chen-Delai-L" }, { "family_name": "Du", "given_name": "Wenbin", "orcid": "0000-0002-7401-1410", "clpid": "Du-Wenbin" }, { "family_name": "Liu", "given_name": "Ying", "clpid": "Liu-Ying" }, { "family_name": "Liu", "given_name": "Weishan", "clpid": "Liu-Weishan" }, { "family_name": "Kuznetsov", "given_name": "Andrey", "clpid": "Kuznetsov-A" }, { "family_name": "Mendez", "given_name": "Felipe E.", "clpid": "Mendez-F-E" }, { "family_name": "Philipson", "given_name": "Louis H.", "clpid": "Philipson-L-H" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Microelectrodes enable localized electrical stimulation and recording, and they have revolutionized our understanding of the spatiotemporal dynamics of systems that generate or respond to\nelectrical signals. However, such comprehensive understanding of\nsystems that rely on molecular signals\u2014e.g., chemical communication in multicellular neural, developmental, or immune systems\u2014remains elusive because of the inability to deliver, capture,\nand interpret complex chemical information. To overcome this\nchallenge, we developed the ''chemistrode,'' a plug-based microfluidic device that enables stimulation, recording, and analysis\nof molecular signals with high spatial and temporal resolution.\nStimulation with and recording of pulses as short as 50 ms was\ndemonstrated. A pair of chemistrodes fabricated by multilayer soft\nlithography recorded independent signals from 2 locations separated by 15 \u03bcm. Like an electrode, the chemistrode does not need\nto be built into an experimental system\u2014it is simply brought into\ncontact with a chemical or biological substrate, and, instead of\nelectrical signals, molecular signals are exchanged. Recorded molecular signals can be injected with additional reagents and analyzed off-line by multiple, independent techniques in parallel (e.g.,\nfluorescence correlation spectroscopy, MALDI-MS, and fluorescence microscopy). When recombined, these analyses provide a\ntime-resolved chemical record of a system's response to stimulation. Insulin secretion from a single murine islet of Langerhans was\nmeasured at a frequency of 0.67 Hz by using the chemistrode. This\narticle characterizes and tests the physical principles that govern\nthe operation of the chemistrode to enable its application to\nprobing local dynamics of chemically responsive matter in chemistry and biology.", "doi": "10.1073/pnas.0807916105", "pmcid": "PMC2579341", "issn": "0027-8424", "publisher": "National Academy of Sciences", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "publication_date": "2008-11-04", "series_number": "44", "volume": "105", "issue": "44", "pages": "16843-16848" }, { "id": "authors:b2j8v-gx577", "collection": "authors", "collection_id": "b2j8v-gx577", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160731095", "type": "article", "title": "Threshold Response of Initiation of Blood Coagulation by Tissue Factor in Patterned Microfluidic Capillaries Is Controlled by Shear Rate", "author": [ { "family_name": "Shen", "given_name": "Feng", "orcid": "0000-0002-4709-330X", "clpid": "Shen-Feng" }, { "family_name": "Kastrup", "given_name": "Christian J.", "clpid": "Kastrup-C-J" }, { "family_name": "Liu", "given_name": "Ying", "clpid": "Liu-Ying" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Objective\u2014Blood flow is considered one of the important parameters that contribute to venous thrombosis. We quantitatively\ntest the relationship between initiation of coagulation and shear rate and suggest a biophysical mechanism to understand\nthis relationship.\nMethods and Results\u2014Flowing human blood and plasma were exposed to cylindrical surfaces patterned with patches of\ntissue factor (TF) by using microfluidics. Initiation of coagulation of normal pooled plasma depended on shear rate, not\nvolumetric flow rate or flow velocity, and coagulation initiated only at shear rates below a critical value. Initiation of\ncoagulation of platelet-rich plasma and whole blood showed similar behavior. At constant shear rate, coagulation of\nplasma also showed a threshold response to the size of a patch of TF, consistent with our previous work in the absence\nof flow.\nConclusion\u2014Initiation of coagulation of flowing blood displays a threshold response to shear rate and to the size of a\nsurface patch of TF. Combined with the results of others, these results set the range of shear rates that limit initiation\nof coagulation by small surface areas of TF and by shear activation of platelets. This range fits the relatively narrow\nrange of physiological shear rates described by Murray's law. (Arterioscler Thromb Vasc Biol. 2008;28:2035-2041)", "doi": "10.1161/ATVBAHA.108.173930", "issn": "1079-5642", "publisher": "American Heart Association", "publication": "Arteriosclerosis, Thrombosis, and Vascular Biology", "publication_date": "2008-11", "series_number": "11", "volume": "28", "issue": "11", "pages": "2035-2041" }, { "id": "authors:ayns4-hc719", "collection": "authors", "collection_id": "ayns4-hc719", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160724242", "type": "article", "title": "Simple Host-Guest Chemistry To Modulate the Process of Concentration and Crystallization of Membrane Proteins by Detergent Capture in a Microfluidic Device", "author": [ { "family_name": "Li", "given_name": "Liang", "clpid": "Li-Liang" }, { "family_name": "Nachtergaele", "given_name": "Sigrid", "clpid": "Nachtergaele-Sigrid" }, { "family_name": "Seddon", "given_name": "Annela M.", "clpid": "Seddon-Annela-M" }, { "family_name": "Tereshko", "given_name": "Valentina", "clpid": "Tereshko-Valentina" }, { "family_name": "Ponomarenko", "given_name": "Nina", "clpid": "Ponomarenko-Nina-S" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper utilizes cyclodextrin-based host-guest chemistry in a microfluidic device to modulate the crystallization of membrane proteins and the process of concentration of membrane protein samples. Methyl-beta-cyclodextrin (MBCD) can efficiently capture a wide variety of detergents commonly used for the stabilization of membrane proteins by sequestering detergent monomers. Reaction Center (RC) from Blastochloris viridis was used here as a model system. In the process of concentrating membrane protein samples, MBCD was shown to break up free detergent micelles and prevent them from being concentrated. The addition of an optimal amount of MBCD to the RC sample captured loosely bound detergent from the protein-detergent complex and improved sample homogeneity, as characterized by dynamic light scattering. Using plug-based microfluidics, RC crystals were grown in the presence of MBCD, giving a different morphology and space group than crystals grown without MBCD. The crystal structure of RC crystallized in the presence of MBCD was consistent with the changes in packing and crystal contacts hypothesized for removal of loosely bound detergent. The incorporation of MBCD into a plug-based microfluidic crystallization method allows efficient use of limited membrane protein sample by reducing the amount of protein required and combining sparse matrix screening and optimization in one experiment. The use of MBCD for detergent capture can be expanded to develop cyclodextrin-derived molecules for fine-tuned detergent capture and thus modulate membrane protein crystallization in an even more controllable way.", "doi": "10.1021/ja805361j", "pmcid": "PMC2596067", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2008-10-29", "series_number": "43", "volume": "130", "issue": "43", "pages": "14324-14328" }, { "id": "authors:20tpx-g8t89", "collection": "authors", "collection_id": "20tpx-g8t89", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160722469", "type": "article", "title": "ABO, D Blood Typing and Subtyping Using Plug-Based Microfluidics", "author": [ { "family_name": "Kline", "given_name": "Timothy R.", "clpid": "Kline-Timothy-R" }, { "family_name": "Runyon", "given_name": "Matthew K.", "clpid": "Runyon-Matthew-K" }, { "family_name": "Pothiawala", "given_name": "Mohammad", "clpid": "Pothiwala-Mohammad" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "A plug-based microfluidic approach was used to perform\nmultiple agglutination assays in parallel without crosscontamination and using only microliter volumes of blood.\nTo perform agglutination assays on-chip, a microfluidic\ndevice was designed to combine aqueous streams of\nantibody, buffer, and red blood cells (RBCs) to form\ndroplets 30-40 nL in volume surrounded by a fluorinated\ncarrier fluid. Using this approach, proof-of-concept ABO\nand D (Rh) blood typing and group A subtyping were\nsuccessfully performed by screening against multiple\nantigens without cross-contamination. On-chip subtyping\ndistinguished common A1 and A2\nRBCs by using a lectinbased dilution assay. This flexible platform was extended\nto differentiate rare, weakly agglutinating RBCs of A\nsubtypes by analyzing agglutination avidity as a function\nof shear rate. Quantitative analysis of changes in contrast\nwithin plugs revealed subtleties in agglutination kinetics\nand enabled characterization of agglutination of rare blood\nsubtypes. Finally, this platform was used to detect bacteria, demonstrating the potential usefulness of this assay\nin detecting sepsis and the potential for applications in\nagglutination-based viral detection. The speed, control,\nand minimal sample consumption provided by this technology present an advance for point of care applications,\nblood typing of newborns, and general blood assays in\nsmall model organisms.", "doi": "10.1021/ac800485q", "pmcid": "PMC2592685", "issn": "0003-2700", "publisher": "American Chemical Society", "publication": "Analytical Chemistry", "publication_date": "2008-08-15", "series_number": "16", "volume": "80", "issue": "16", "pages": "6190-6197" }, { "id": "authors:e9mf6-5tr54", "collection": "authors", "collection_id": "e9mf6-5tr54", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160729017", "type": "article", "title": "Rate of mixing controls rate and outcome of autocatalytic processes\u2014theory and microfluidic experiments with chemical reactions and blood coagulation", "author": [ { "family_name": "Pompano", "given_name": "Rebecca R.", "clpid": "Pompano-Rebecca-R" }, { "family_name": "Li", "given_name": "Hung-Wing", "clpid": "Li-Hung-Wing" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This article demonstrates that the rate of mixing can regulate the rate and outcome of both biological and\nnonbiological autocatalytic reaction systems that display a threshold response to the concentration of an activator. Plug-based\nmicrofluidics was used to control the timing of reactions, the rate of mixing, and surface chemistry in blood clotting and its\nchemical model. Initiation of clotting of human blood plasma required addition of a critical concentration of thrombin. Clotting\ncould be prevented by rapid mixing when thrombin was added near the critical concentration, and mixing also affected the rate\nof clotting when thrombin was added at concentrations far above the critical concentration in two clinical clotting assays for\nhuman plasma. This phenomenon was modeled by a simple mechanism\u2014local and global competition between the clotting\nreaction, which autocatalytically produces an activator, and mixing, which removes the activator. Numerical simulations showed\nthat the Damk\u00f6hler number, which describes this competition, predicts the effects of mixing. Many biological systems are controlled by thresholds, and these results shed light on the dynamics of these systems in the presence of spatial heterogeneities\nand provide simple guidelines for designing and interpreting experiments with such systems.", "doi": "10.1529/biophysj.108.129486", "pmcid": "PMC2479617", "issn": "0006-3495", "publisher": "Biophysical Society", "publication": "Biophysical Journal", "publication_date": "2008-08-01", "series_number": "3", "volume": "95", "issue": "3", "pages": "1531-1543" }, { "id": "authors:rqc6r-mj822", "collection": "authors", "collection_id": "rqc6r-mj822", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160721104", "type": "article", "title": "Using chemistry and microfluidics to understand the spatial dynamics of complex biological networks", "author": [ { "family_name": "Kastrup", "given_name": "Christian J.", "clpid": "Kastrup-Christian-J" }, { "family_name": "Runyon", "given_name": "Matthew K.", "clpid": "Runyon-Matthew-K" }, { "family_name": "Lucchetta", "given_name": "Elena M.", "clpid": "Lucchetta-Elena-M" }, { "family_name": "Price", "given_name": "Jessica M.", "clpid": "Price-Jessica-M" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Understanding the spatial dynamics of biochemical networks is both fundamentally important for understanding life at the systems level and also has practical implications for medicine, engineering, biology, and chemistry. Studies at the level of individual reactions provide essential information about the function, interactions, and localization of individual molecular species and reactions in a network. However, analyzing the spatial dynamics of complex biochemical networks at this level is difficult. Biochemical networks are non-equilibrium systems containing dozens to hundreds of reactions with nonlinear and time-dependent interactions, and these interactions are influenced by diffusion, flow, and the relative values of state-dependent kinetic parameters. To achieve an overall understanding of the spatial dynamics of a network and the global mechanisms that drive its function, networks must be analyzed as a whole, where all of the components and influential parameters of a network are simultaneously considered. Here, we describe chemical concepts and microfluidic tools developed for network-level investigations of the spatial dynamics of these networks. Modular approaches can be used to simplify these networks by separating them into modules, and simple experimental or computational models can be created by replacing each module with a single reaction. Microfluidics can be used to implement these models as well as to analyze and perturb the complex network itself with spatial control on the micrometer scale. We also describe the application of these network-level approaches to elucidate the mechanisms governing the spatial dynamics of two networks-hemostasis (blood clotting) and early patterning of the Drosophila embryo. To investigate the dynamics of the complex network of hemostasis, we simplified the network by using a modular mechanism and created a chemical model based on this mechanism by using microfluidics. Then, we used the mechanism and the model to predict the dynamics of initiation and propagation of blood clotting and tested these predictions with human blood plasma by using microfluidics. We discovered that both initiation and propagation of clotting are regulated by a threshold response to the concentration of activators of clotting, and that clotting is sensitive to the spatial localization of stimuli. To understand the dynamics of patterning of the Drosophila embryo, we used microfluidics to perturb the environment around a developing embryo and observe the effects of this perturbation on the expression of Hunchback, a protein whose localization is essential to proper development. We found that the mechanism that is responsible for Hunchback positioning is asymmetric, time-dependent, and more complex than previously proposed by studies of individual reactions. Overall, these approaches provide strategies for simplifying, modeling, and probing complex networks without sacrificing the functionality of the network. Such network-level strategies may be most useful for understanding systems with non-linear interactions where spatial dynamics is essential for function. In addition, microfluidics provides an opportunity to investigate the mechanisms responsible for robust functioning of complex networks. By creating nonideal, stressful, and perturbed environments, microfluidic experiments could reveal the function of pathways thought to be nonessential under ideal conditions.", "doi": "10.1021/ar700174g", "pmcid": "PMC2593841", "issn": "0001-4842", "publisher": "American Chemical Society", "publication": "Accounts of Chemical Research", "publication_date": "2008-04-15", "series_number": "4", "volume": "41", "issue": "4", "pages": "549-558" }, { "id": "authors:hz44t-esr26", "collection": "authors", "collection_id": "hz44t-esr26", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160730150", "type": "article", "title": "Effects of shear rate on propagation of blood clotting determined using microfluidics and numerical simulations", "author": [ { "family_name": "Runyon", "given_name": "Matthew K.", "clpid": "Runyon-M-K" }, { "family_name": "Kastrup", "given_name": "Christian J.", "clpid": "Kastrup-C-J" }, { "family_name": "Johnson-Kerner", "given_name": "Bethany L.", "clpid": "Johnson-Kerner-B-L" }, { "family_name": "Van Ha", "given_name": "Thuong G.", "clpid": "Van-Ha-Thuong-G" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper describes microfluidic experiments with human blood plasma and numerical simulations to determine the role of fluid flow in the regulation of propagation of blood clotting. We demonstrate that propagation of clotting can be regulated by different mechanisms depending on the volume-to-surface ratio of a channel. In small channels, propagation of clotting can be prevented by surface-bound inhibitors of clotting present on vessel walls. In large channels, where surface-bound inhibitors are ineffective, propagation of clotting can be prevented by a shear rate above a threshold value, in agreement with predictions of a simple reaction-diffusion mechanism. We also demonstrate that propagation of clotting in a channel with a large volume-to-surface ratio and a shear rate below a threshold shear rate can be slowed by decreasing the production of thrombin, an activator of clotting. These in vitro results make two predictions, which should be experimentally tested in vivo. First, propagation of clotting from superficial veins to deep veins may be regulated by shear rate, which might explain the correlation between superficial thrombosis and the development of deep vein thrombosis (DVT). Second, nontoxic thrombin inhibitors with high binding affinities could be locally administered to prevent recurrent thrombosis after a clot has been removed. In addition, these results demonstrate the utility of simplified mechanisms and microfluidics for generating and testing predictions about the dynamics of complex biochemical networks.", "doi": "10.1021/ja076301r", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2008-03-19", "series_number": "11", "volume": "130", "issue": "11", "pages": "3458-3464" }, { "id": "authors:8eteb-x0k54", "collection": "authors", "collection_id": "8eteb-x0k54", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160716197", "type": "article", "title": "Detecting bacteria and determining their susceptibility to antibiotics by stochastic confinement in nanoliter droplets using plug-based microfluidics", "author": [ { "family_name": "Boedicker", "given_name": "James Q.", "orcid": "0000-0003-4107-3719", "clpid": "Boedicker-James-Q" }, { "family_name": "Li", "given_name": "Liang", "clpid": "Li-Liang" }, { "family_name": "Kline", "given_name": "Timothy R.", "clpid": "Kline-Timothy-R" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This article describes plug-based microfluidic technology that enables rapid detection and drug\nsusceptibility screening of bacteria in samples, including complex biological matrices, without preincubation. Unlike conventional bacterial culture and detection methods, which rely on incubation\nof a sample to increase the concentration of bacteria to detectable levels, this method confines individual bacteria into droplets nanoliters in volume. When single cells are confined into plugs of small\nvolume such that the loading is less than one bacterium per plug, the detection time is proportional\nto plug volume. Confinement increases cell density and allows released molecules to accumulate\naround the cell, eliminating the pre-incubation step and reducing the time required to detect\nthe bacteria. We refer to this approach as 'stochastic confinement'. Using the microfluidic hybrid\nmethod, this technology was used to determine the antibiogram \u2013 or chart of antibiotic sensitivity\n\u2013 of methicillin-resistant Staphylococcus aureus (MRSA) to many antibiotics in a single experiment\nand to measure the minimal inhibitory concentration (MIC) of the drug cefoxitin (CFX) against\nthis strain. In addition, this technology was used to distinguish between sensitive and resistant\nstrains of S. aureus in samples of human blood plasma. High-throughput microfluidic techniques\ncombined with single-cell measurements also enable multiple tests to be performed simultaneously\non a single sample containing bacteria. This technology may provide a method of rapid\nand effective patient-specific treatment of bacterial infections and could be extended to a variety\nof applications that require multiple functional tests of bacterial samples on reduced timescales.", "doi": "10.1039/B804911D", "pmcid": "PMC2612531", "issn": "1473-0197", "publisher": "Royal Society of Chemistry", "publication": "Lab on a Chip", "publication_date": "2008", "series_number": "8", "volume": "8", "issue": "8", "pages": "1265-1272" }, { "id": "authors:v0v0z-cm610", "collection": "authors", "collection_id": "v0v0z-cm610", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20141212-131800389", "type": "article", "title": "Using microfluidics to understand the effect of spatial distribution of tissue factor on blood coagulation", "author": [ { "family_name": "Shen", "given_name": "Feng", "orcid": "0000-0002-4709-330X", "clpid": "Shen-Feng" }, { "family_name": "Kastrup", "given_name": "Christian J.", "clpid": "Kastrup-C-J" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Initiation of blood coagulation by tissue factor (TF) is a robust, highly regulated process. Both the spatial distribution of TF and the geometry of the vasculature may play important roles in regulating coagulation. As this review describes, microfluidic systems provide a unique opportunity for investigating the spatiotemporal dynamics of blood coagulation in vitro. Microfluidic systems with surfaces of phospholipid bilayers patterned with TF have been used to demonstrate experimentally the threshold responses of initiation of coagulation to the size and shape of surfaces presenting TF. These systems have also been used to demonstrate experimentally that propagation of coagulation is regulated by the shear rate of blood flow in microcapillaries and microchannels. By understanding these and other aspects of the spatial dynamics that regulate blood coagulation, many new methods for treating clotting disorders, such as venous thromboembolism (VTE) and sepsis, could arise.", "doi": "10.1016/S0049-3848(08)70015-X", "issn": "0049-3848", "publisher": "Elsevier", "publication": "Thrombosis Research", "publication_date": "2008", "series_number": "Suppl. 1", "volume": "122", "issue": "Suppl. 1", "pages": "S27-S30" }, { "id": "authors:63m5d-r9p84", "collection": "authors", "collection_id": "63m5d-r9p84", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160719738", "type": "article", "title": "Can we build synthetic, multicellular systems by controlling developmental signaling in space and time?", "author": [ { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Maharbiz", "given_name": "Michel M.", "clpid": "Maharbiz-Michel-M" } ], "abstract": "Using biological machinery to make new, functional molecules\nis an exciting area in chemical biology. Complex molecules\ncontaining both 'natural' and 'unnatural' components are made\nby processes ranging from enzymatic catalysis to the\ncombination of molecular biology with chemical tools. Here, we\ndiscuss applying this approach to the next level of biological\ncomplexity \u2014 building synthetic, functional biotic systems by\nmanipulating biological machinery responsible for\ndevelopment of multicellular organisms. We describe recent\nadvances enabling this approach, including first, recent\ndevelopmental biology progress unraveling the pathways and\nmolecules involved in development and pattern formation;\nsecond, emergence of microfluidic tools for delivering stimuli to\na developing organism with exceptional control in space and\ntime; third, the development of molecular and synthetic biology\ntoolsets for redesigning or de novo engineering of signaling\nnetworks; and fourth, biological systems that are especially\namendable to this approach.", "doi": "10.1016/j.cbpa.2007.10.003", "pmcid": "PMC2203304", "issn": "1367-5931", "publisher": "Elsevier", "publication": "Current Opinion in Chemical Biology", "publication_date": "2007-12", "series_number": "6", "volume": "11", "issue": "6", "pages": "604-611" }, { "id": "authors:xbvf0-pth52", "collection": "authors", "collection_id": "xbvf0-pth52", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160721703", "type": "article", "title": "Characterization of the Threshold Response of Initiation of Blood Clotting to Stimulus Patch Size", "author": [ { "family_name": "Kastrup", "given_name": "Christian J.", "clpid": "Kastrup-Christian-J" }, { "family_name": "Shen", "given_name": "Feng", "orcid": "0000-0002-4709-330X", "clpid": "Shen-Feng" }, { "family_name": "Runyon", "given_name": "Matthew K.", "clpid": "Runyon-Matthew-K" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This article demonstrates that the threshold response of initiation of blood clotting to the size of a patch of stimulus is a robust phenomenon under a wide range of conditions and follows a simple scaling relationship based on the Damkohler number. Human blood and plasma were exposed to surfaces patterned with patches presenting clotting stimuli using microfluidics. Perturbations of the complex network of hemostasis, including temperature, variations in the concentration of stimulus (tissue factor), and the absence or inhibition of individual components of the network (factor IIa, factor V, factor VIII, and thrombomodulin), did not affect the existence of this response. A scaling relationship between the threshold patch size and the timescale of reaction for clotting was supported in numerical simulations, a simple chemical model system, and experiments with human blood plasma. These results may be useful for understanding the spatiotemporal dynamics of other autocatalytic systems and emphasize the relevance of clustering of proteins and lipids in the regulation of signaling processes.", "doi": "10.1529/biophysj.107.109009", "pmcid": "PMC1989713", "issn": "0006-3495", "publisher": "Biophysical Society", "publication": "Biophysical Journal", "publication_date": "2007-10-15", "series_number": "8", "volume": "93", "issue": "8", "pages": "2969-2977" }, { "id": "authors:e4xjv-wpy69", "collection": "authors", "collection_id": "e4xjv-wpy69", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160720940", "type": "article", "title": "A physical organic mechanistic approach to understanding the complex reaction network of hemostasis (blood clotting)", "author": [ { "family_name": "Kastrup", "given_name": "Christian J.", "clpid": "Kastrup-C-J" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This review focuses on how the mechanistic approach of physical organic chemistry can be used to elucidate the mechanisms behind complex biochemical networks. The dynamics of biochemical reaction networks is difficult to describe by considering their individual reactions, just as the dynamics of organic reactions is difficult to describe by considering individual electrons and atomic nuclei. Physical organic chemists have developed a useful set of tools to predict the outcome of organic reactions by separating the interacting molecules into modules (functional groups), and defining general rules for how these modules interact (mechanisms). This review shows how these tools of physical organic chemistry may be used to describe reaction networks. In addition, it describes the application of these tools to develop a mechanistic understanding of the dynamics of the complex network of hemostasis, which regulates blood clotting.", "doi": "10.1002/poc.1242", "issn": "0894-3230", "publisher": "Wiley", "publication": "Journal of Physical Organic Chemistry", "publication_date": "2007-10", "series_number": "10", "volume": "20", "issue": "10", "pages": "711-715" }, { "id": "authors:c32j4-xft93", "collection": "authors", "collection_id": "c32j4-xft93", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160728859", "type": "article", "title": "Attachment of cells to islands presenting gradients of adhesion ligands", "author": [ { "family_name": "Petty", "given_name": "Rafe T.", "clpid": "Petty-Rafe-T" }, { "family_name": "Li", "given_name": "Hung-Wing", "clpid": "Li-Hung-Wing" }, { "family_name": "Maduram", "given_name": "Jane H.", "clpid": "Maduram-Jane-H" }, { "family_name": "Ismagilov", "given_name": "Rustem", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Mrksich", "given_name": "Milan", "orcid": "0000-0002-4964-796X", "clpid": "Mrksich-Milan" } ], "abstract": "This paper reports a strategy that uses microfluidic networks to pattern self-assembled monolayers with gradient microislands for the attachment of individual cells. A microfluidic network is first used to pattern a monolayer into square regions that present maleimide groups and then used to flow a solution having a gradient of the cell adhesion peptide Arg-Gly-Asp over the substrate. In this way, the surface is patterned with microislands approximately 33 x 33 micrometers in size and each having a defined gradient of immobilized cell adhesion ligand. B16F10 cells were allowed to attach to the patterned islands and were found to display a nonuniform distribution of cytoskeletal structures in response to the gradient of adhesion ligand. This work is significant because it permits studies of the influence of a nonuniform microenvironment on the polarization, differentiation, and signaling of adherent cells.", "doi": "10.1021/ja0735709", "pmcid": "PMC2543034", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2007-07-25", "series_number": "29", "volume": "129", "issue": "29", "pages": "8966-8967" }, { "id": "authors:sjn7h-bv729", "collection": "authors", "collection_id": "sjn7h-bv729", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160729975", "type": "article", "title": "Propagation of blood clotting in the complex biochemical network of hemostasis is described by a simple mechanism", "author": [ { "family_name": "Runyon", "given_name": "Matthew K.", "clpid": "Runyon-M-K" }, { "family_name": "Johnson-Kerner", "given_name": "Bethany L.", "clpid": "Johnson-Kerner-B-L" }, { "family_name": "Kastrup", "given_name": "Christian J.", "clpid": "Kastrup-C-J" }, { "family_name": "Van Ha", "given_name": "Thuong G.", "clpid": "Van-Ha-Thuong-G" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Hemostasis is the complex biochemical network that controls blood clotting. We previously described a chemical model that mimicked the dynamics of hemostasis based on a simple regulatory mechanisma threshold response due to the competition between production and removal of activators. Here, we used human blood plasma in phospholipid-coated microfluidic channels to test predictions based on this mechanism. We demonstrated that, for a given geometry of channels, clot propagation from an obstructed channel into a channel with flowing blood plasma is dependent on the shear rate in the channel with flowing blood plasma. If confirmed in vivo, these results may explain clot propagation from a small vessel to a larger, clinically relevant vessel. In addition, these results would further validate the use of modular mechanisms, simplified chemical models, and microfluidics to study complex biochemical networks.", "doi": "10.1021/ja072602p", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2007-06-06", "series_number": "22", "volume": "129", "issue": "22", "pages": "7014-7015" }, { "id": "authors:9fv09-b9f68", "collection": "authors", "collection_id": "9fv09-b9f68", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160721515", "type": "article", "title": "Response to shape emerges in a complex biochemical network and its simple chemical analogue", "author": [ { "family_name": "Kastrup", "given_name": "Christian J.", "clpid": "Kastrup-C-J" }, { "family_name": "Shen", "given_name": "Feng", "orcid": "0000-0002-4709-330X", "clpid": "Shen-Feng" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "n/a", "doi": "10.1002/anie.200604995", "issn": "1433-7851", "publisher": "Wiley", "publication": "Angewandte Chemie International Edition", "publication_date": "2007-05-11", "series_number": "20", "volume": "46", "issue": "20", "pages": "3660-3662" }, { "id": "authors:46gj7-60s97", "collection": "authors", "collection_id": "46gj7-60s97", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160723117", "type": "article", "title": "Using a Multijunction Microfluidic Device To Inject Substrate into an Array of Preformed Plugs without Cross-Contamination: Comparing Theory and Experiments", "author": [ { "family_name": "Li", "given_name": "Liang", "clpid": "Li-Liang" }, { "family_name": "Boedicker", "given_name": "James Q.", "orcid": "0000-0003-4107-3719", "clpid": "Boedicker-James-Q" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Abstract: In this paper we describe a multijunction microfluidic device for the injection of a substrate into an array of preformed plugs carried by an immiscible fluid in a microchannel. The device uses multiple junctions to inject substrate into preformed plugs without synchronization of the flow of substrate and the array of preformed plugs of reagent, which reduces cross-contamination of the plugs, eliminates formation of small droplets of substrate, and allows a greater range of injection ratios compared to that of a single T-junction. The device was based on a previously developed physical model for transport that was here adapted to describe injection and experimentally verified. After characterization, the device was applied to two biochemical assays, including evaluation of the enzymatic activity of thrombin and determination of the coagulation time of human blood plasma, which both provided reliable results. The reduction of cross-contamination and greater range of injection ratios achieved by this device may improve the processes that involve addition and titration of reagents into plugs, such as high-throughput screening of protein crystallization conditions.", "doi": "10.1021/ac062179n", "pmcid": "PMC2080796", "issn": "0003-2700", "publisher": "American Chemical Society", "publication": "Analytical Chemistry", "publication_date": "2007-04-01", "series_number": "7", "volume": "79", "issue": "7", "pages": "2756-2761" }, { "id": "authors:wpdey-73g81", "collection": "authors", "collection_id": "wpdey-73g81", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160717523", "type": "article", "title": "Using three-phase flow of immiscible liquids to prevent coalescence of droplets in microfluidic channels: Criteria to identify the third liquid and validation with protein crystallization", "author": [ { "family_name": "Chen", "given_name": "Delai L.", "clpid": "Chen-Delai-L" }, { "family_name": "Li", "given_name": "Liang", "clpid": "Li-Liang" }, { "family_name": "Reyes", "given_name": "Sebastian", "clpid": "Reyes-Sebastian" }, { "family_name": "Adamson", "given_name": "David N.", "clpid": "Adamson-David-N" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This manuscript describes the effect of interfacial tensions on three-phase liquid-liquid-liquid flow in microfluidic channels and the use of this flow to prevent microfluidic plugs from coalescing. One problem in using microfluidic plugs as microreactors is the coalescence of adjacent plugs caused by the relative motion of plugs during flow. Here, coalescence of reagent plugs was eliminated by using plugs of a third immiscible liquid as spacers to separate adjacent reagent plugs. This work tested the requirements of interfacial tensions for plugs of a third liquid to be effective spacers. Two candidates satisfying the requirements were identified, and one of these liquids was used in the crystallization of protein human Tdp1 to demonstrate its compatibility with protein crystallization in plugs. This method for identifying immiscible liquids for use as a spacer will also be useful for applications involving manipulation of large arrays of droplets in microfluidic channels.", "doi": "10.1021/la062152z", "pmcid": "PMC1986632", "issn": "0743-7463", "publisher": "American Chemical Society", "publication": "Langmuir", "publication_date": "2007-02-13", "series_number": "4", "volume": "23", "issue": "4", "pages": "2255-2260" }, { "id": "authors:d06ze-1dq58", "collection": "authors", "collection_id": "d06ze-1dq58", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160724073", "type": "article", "title": "Nanoliter microfluidic hybrid method for simultaneous screening and optimization validated with crystallization of membrane proteins", "author": [ { "family_name": "Li", "given_name": "Liang", "clpid": "Li-Liang" }, { "family_name": "Mustafi", "given_name": "Debarshi", "clpid": "Mustafi-D" }, { "family_name": "Fu", "given_name": "Qiang", "clpid": "Fu-Qiang" }, { "family_name": "Tereshko", "given_name": "Valentina", "clpid": "Tereshko-V" }, { "family_name": "Chen", "given_name": "Delai L.", "clpid": "Chen-Delai-L" }, { "family_name": "Tice", "given_name": "Joshua D.", "clpid": "Tice-J-D" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "High-throughput screening and optimization experiments are critical to a number of fields, including chemistry and structural and molecular biology. The separation of these two steps may introduce false negatives and a time delay between initial screening and subsequent optimization. Although a hybrid method combining both steps may address these problems, miniaturization is required to minimize sample consumption. This article reports a \"hybrid\" droplet-based microfluidic approach that combines the steps of screening and optimization into one simple experiment and uses nanoliter-sized plugs to minimize sample consumption. Many distinct reagents were sequentially introduced as approx. 140-nl plugs into a microfluidic device and combined with a substrate and a diluting buffer. Tests were conducted in approx. 10-nl plugs containing different concentrations of a reagent. Methods were developed to form plugs of controlled concentrations, index concentrations, and incubate thousands of plugs inexpensively and without evaporation. To validate the hybrid method and demonstrate its applicability to challenging problems, crystallization of model membrane proteins and handling of solutions of detergents and viscous precipitants were demonstrated. By using 10 \u03bcl of protein solution, approx. 1,300 crystallization trials were set up within 20 min by one researcher. This method was compatible with growth, manipulation, and extraction of high-quality crystals of membrane proteins, demonstrated by obtaining high-resolution diffraction images and solving a crystal structure. This robust method requires inexpensive equipment and supplies, should be especially suitable for use in individual laboratories, and could find applications in a number of areas that require chemical, biochemical, and biological screening and optimization.", "doi": "10.1073/pnas.0607502103", "pmcid": "PMC1748211", "issn": "0027-8424", "publisher": "National Academy of Sciences", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "publication_date": "2006-12-19", "series_number": "51", "volume": "103", "issue": "51", "pages": "19243-19248" }, { "id": "authors:5ft01-pwm48", "collection": "authors", "collection_id": "5ft01-pwm48", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160718311", "type": "article", "title": "Time-Controlled Microfluidic Seeding in nL-Volume Droplets To Separate Nucleation and Growth Stages of Protein Crystallization", "author": [ { "family_name": "Gerdts", "given_name": "Cory J.", "clpid": "Gerdts-Cory-J" }, { "family_name": "Tereshko", "given_name": "Valentina", "clpid": "Tereshko-Valentina" }, { "family_name": "Yadav", "given_name": "Maneesh K.", "clpid": "Yadav-Maneesh-K" }, { "family_name": "Dementieva", "given_name": "Irina", "clpid": "Dementieva-Irina" }, { "family_name": "Collart", "given_name": "Frank", "clpid": "Collart-Frank" }, { "family_name": "Joachimiak", "given_name": "Andrzej", "clpid": "Joachimiak-Andrzej" }, { "family_name": "Stevens", "given_name": "Raymond C.", "orcid": "0000-0002-4522-8725", "clpid": "Stevens-Raymond-C" }, { "family_name": "Kuhn", "given_name": "Peter", "clpid": "Kuhn-Peter" }, { "family_name": "Kossiakoff", "given_name": "Anthony", "orcid": "0000-0003-3174-9359", "clpid": "Kossiakoff-Anthony-A" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper describes a method of time-controlled seeding to separate the stages of nucleation and growth in protein crystallization using a microfluidic device.", "doi": "10.1002/anie.200602946", "pmcid": "PMC1766323", "issn": "1433-7851", "publisher": "Wiley", "publication": "Angewandte Chemie International Edition", "publication_date": "2006-12-11", "series_number": "48", "volume": "45", "issue": "48", "pages": "8156-8160" }, { "id": "authors:p9d6v-w7z37", "collection": "authors", "collection_id": "p9d6v-w7z37", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160732141", "type": "article", "title": "Reactions in droplets in microfluidic channels", "author": [ { "family_name": "Song", "given_name": "Helen", "clpid": "Song-Helen" }, { "family_name": "Chen", "given_name": "Delai L.", "clpid": "Chen-Delai-L" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Fundamental and applied research in chemistry and biology benefits\nfrom opportunities provided by droplet-based microfluidic systems.\nThese systems enable the miniaturization of reactions by compartmentalizing reactions in droplets of femoliter to microliter volumes.\nCompartmentalization in droplets provides rapid mixing of reagents,\ncontrol of the timing of reactions on timescales from milliseconds to\nmonths, control of interfacial properties, and the ability to synthesize\nand transport solid reagents and products. Droplet-based microfluidics can help to enhance and accelerate chemical and biochemical\nscreening, protein crystallization, enzymatic kinetics, and assays.\nMoreover, the control provided by droplets in microfluidic devices can\nlead to new scientific methods and insights.", "doi": "10.1002/anie.200601554", "pmcid": "PMC1766322", "issn": "1433-7851", "publisher": "Wiley", "publication": "Angewandte Chemie International Edition", "publication_date": "2006-11-13", "series_number": "44", "volume": "45", "issue": "44", "pages": "7336-7356" }, { "id": "authors:xwgv0-s3q82", "collection": "authors", "collection_id": "xwgv0-s3q82", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160721372", "type": "article", "title": "Modular chemical mechanism predicts spatiotemporal dynamics of initiation in the complex network of hemostasis", "author": [ { "family_name": "Kastrup", "given_name": "Christian J.", "clpid": "Kastrup-C-J" }, { "family_name": "Runyon", "given_name": "Matthew K.", "clpid": "Runyon-M-K" }, { "family_name": "Shen", "given_name": "Feng", "orcid": "0000-0002-4709-330X", "clpid": "Shen-Feng" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This article demonstrates that a simple chemical model system, built by using a modular approach, may be used to predict the spatiotemporal dynamics of initiation of blood clotting in the complex network of hemostasis. Microfluidics was used to create in vitro environments that expose both the complex network and the model system to surfaces patterned with patches presenting clotting stimuli. Both systems displayed a threshold response, with clotting initiating only on isolated patches larger than a threshold size. The magnitude of the threshold patch size for both systems was described by the Damkohler number, measuring competition of reaction and diffusion. Reaction produces activators at the patch, and diffusion removes activators from the patch. The chemical model made additional predictions that were validated experimentally with human blood plasma. These experiments show that blood can be exposed to significant amounts of clot-inducing stimuli, such as tissue factor, without initiating clotting. Overall, these results demonstrate that such chemical model systems, implemented with microfluidics, may be used to predict spatiotemporal dynamics of complex biochemical networks.", "doi": "10.1073/pnas.0605560103", "pmcid": "PMC1635074", "issn": "0027-8424", "publisher": "National Academy of Sciences", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "publication_date": "2006-10-24", "series_number": "43", "volume": "103", "issue": "43", "pages": "15747-15752" }, { "id": "authors:w9zwk-d1t14", "collection": "authors", "collection_id": "w9zwk-d1t14", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160732540", "type": "article", "title": "On-chip titration of an anticoagulant argatroban and determination of the clotting time within whole blood or plasma using a plug-based microfluidic system", "author": [ { "family_name": "Song", "given_name": "Helen", "clpid": "Song-Helen" }, { "family_name": "Li", "given_name": "Hung-Wing", "clpid": "Li-Hung-Wing" }, { "family_name": "Munson", "given_name": "Matthew S.", "clpid": "Munson-Matthew-S" }, { "family_name": "Van Ha", "given_name": "Thuong G.", "clpid": "Van-Ha-Thuong-G" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper describes extending plug-based microfluidics to handling complex biological fluids such as blood, solving the problem of injecting additional reagents into plugs, and applying this system to measuring of clotting time in small volumes of whole blood and plasma. Plugs are droplets transported through microchannels by fluorocarbon fluids. A plug-based microfluidic system was developed to titrate an anticoagulant (argatroban) into blood samples and to measure the clotting time using the activated partial thromboplastin time (APTT) test. To carry out these experiments, the following techniques were developed for a plug-based system: (i) using Teflon AF coating on the microchannel wall to enable formation of plugs containing blood and transport of the solid fibrin clots within plugs, (ii) using a hydrophilic glass capillary to enable reliable merging of a reagent from an aqueous stream into plugs, (iii) using bright-field microscopy to detect the formation of a fibrin clot within plugs and using fluorescent microscopy to detect the production of thrombin using a fluorogenic substrate, and (iv) titration of argatroban (0-1.5 mu g/mL) into plugs and measurement of the resulting APTTs at room temperature (23 degrees C) and physiological temperature (37 degrees C). APTT measurements were conducted with normal pooled plasma (platelet-poor plasma) and with donor's blood samples ( both whole blood and platelet-rich plasma). APTT values and APTT ratios measured by the plug-based microfluidic device were compared to the results from a clinical laboratory at 37 degrees C. APTT obtained from the on-chip assay were about double those from the clinical laboratory but the APTT ratios from these two methods agreed well with each other.", "doi": "10.1021/ac0601718", "pmcid": "PMC1851927", "issn": "0003-2700", "publisher": "American Chemical Society", "publication": "Analytical Chemistry", "publication_date": "2006-07-15", "series_number": "14", "volume": "78", "issue": "14", "pages": "4839-4849" }, { "id": "authors:ss744-gq430", "collection": "authors", "collection_id": "ss744-gq430", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160717350", "type": "article", "title": "Microfluidic cartridges preloaded with nanoliter plugs of reagents: an alternative to 96-well plates for screening", "author": [ { "family_name": "Chen", "given_name": "Delai L.", "clpid": "Chen-Delai-L" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "In traditional screening with 96-well plates, microliters of substrates are consumed for each reaction. Further miniaturization is limited by the special equipment and techniques required to dispense nanoliter volumes of fluid. Plug-based microfluidics confines reagents in nanoliter plugs (droplets surrounded by fluorinated carrier fluid), and uses simple pumps to control the flow of plugs. By using cartridges pre-loaded with nanoliter plugs of reagents, only two pumps and a merging junction are needed to set up a screen. Screening with preloaded cartridges uses only nanoliters of substrate per reaction, and requires no microfabrication. The low cost and simplicity of this method has the potential of replacing 96-well and other multi-well plates, and has been applied to enzymatic assays, protein crystallization and optimization of organic reactions.", "doi": "10.1016/j.cbpa.2006.04.004", "pmcid": "PMC1764868", "issn": "1367-5931", "publisher": "Elsevier", "publication": "Current Opinion in Chemical Biology", "publication_date": "2006-06", "series_number": "3", "volume": "10", "issue": "3", "pages": "226-231" }, { "id": "authors:sw90n-wh349", "collection": "authors", "collection_id": "sw90n-wh349", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160718478", "type": "article", "title": "Microgram-scale testing of reaction conditions in solution using nanoliter plugs in microfluidics with detection by MALDI-MS", "author": [ { "family_name": "Hatakeyama", "given_name": "Takuji", "clpid": "Hatakeyama-Takuji" }, { "family_name": "Chen", "given_name": "Delai L.", "clpid": "Chen-Delai-L" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper describes a microfluidic system to screen and optimize organic reaction conditions on a submicrogram scale. The system uses discrete droplets (plugs) as microreactors separated and transported by a continuous phase of a fluorinated carrier fluid. Previously, we demonstrated the use of a microfabricated PDMS plug-based microfluidic system to perform assays and crystallization experiments in aqueous solutions with optical detection. Here, we developed an approach that does not require microfabrication of microfluidic devices, is applicable to synthetic reactions in organic solvents, and uses detection by MALDI-MS. As a demonstration, conditions for selective deacetylation of ouabain hexaacetate were tested, and the optimum conditions for mono-, bis-, or trisdeacetylation have been identified. These conditions were validated by scale-up reactions and isolating these potentially neurotoxic products. Mono- and bisdeacetylated products are unstable intermediates in the deacetylation and were isolated for the first time. This system enables no-loss handling of submicroliter volumes containing a few micrograms of a compound of interest. It could become valuable for investigating or optimizing reactions of precious substrates (e.g., products of long synthetic sequences and natural products that can be isolated only in small quantities).", "doi": "10.1021/ja057720w", "pmcid": "PMC1851926", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2006-03-01", "series_number": "8", "volume": "128", "issue": "8", "pages": "2518-2519" }, { "id": "authors:ykjg2-md758", "collection": "authors", "collection_id": "ykjg2-md758", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160725195", "type": "article", "title": "Characterization of the local temperature in space and time around a developing Drosophila embryo in a microfluidic device", "author": [ { "family_name": "Lucchetta", "given_name": "Elena M.", "clpid": "Lucchetta-E-M" }, { "family_name": "Munson", "given_name": "Matthew S.", "clpid": "Munson-M-S" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper characterizes a microfluidic platform that differentially controls the temperature of each half of a living Drosophila melanogaster fruitfly embryo in space and time (E. M. Lucchetta, J. H. Lee, L. A. Fu, N. H. Patel and R. F. Ismagilov, Nature, 2005, 434, 1134-1138). This platform relies on laminar flow of two streams of liquid with different temperature, and on rapid prototyping in polydimethylsiloxane (PDMS). Here, we characterized fluid flow and heat transport in this platform both experimentally and by numerical simulation, and estimated the temperature distribution around and within the embryo by numerical simulation, to identify the conditions for creating a sharper temperature difference (temperature step) over the embryo. Embryos were removed from the device and immunostained histochemically for detection of Paired protein. Biochemical processes are sensitive to small differences in environmental temperature. The microfluidic platform characterized here could prove useful in understanding dynamics of biochemical networks as they respond to changes in temperature.", "doi": "10.1039/B516119C", "issn": "1473-0197", "publisher": "Royal Society of Chemistry", "publication": "Lab on a Chip", "publication_date": "2006", "series_number": "2", "volume": "6", "issue": "2", "pages": "185-190" }, { "id": "authors:9wybq-h6d37", "collection": "authors", "collection_id": "9wybq-h6d37", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160715778", "type": "article", "title": "Production of arrays of chemically distinct nanolitre plugs via repeated splitting in microfluidic devices", "author": [ { "family_name": "Adamson", "given_name": "David N.", "clpid": "Adamson-David-N" }, { "family_name": "Mustafi", "given_name": "Debarshi", "clpid": "Mustafi-Debarshi" }, { "family_name": "Zhang", "given_name": "John X. J.", "clpid": "Zhang-John-X-J" }, { "family_name": "Zheng", "given_name": "Bo", "orcid": "0000-0001-8344-3445", "clpid": "Zheng-Bo" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper reports a method for the production of arrays of nanolitre plugs with distinct chemical compositions. One of the primary constraints on the use of plug-based microfluidics for large scale biological screening is the difficulty of fabricating arrays of chemically distinct plugs on the nanolitre scale. Here, using microfluidic devices with several T-junctions linked in series, a single input array of large (similar to 320 nL) plugs was split to produce 16 output arrays of smaller (similar to 20 nL) plugs; the composition and configuration of these arrays were identical to that of the input. This paper shows how the passive break-up of plugs in T-junction microchannel geometries can be used to produce a set of smaller-volume output arrays useful for chemical screening from a single large-volume array. A simple theoretical description is presented to describe splitting as a function of the Capillary number, the capillary pressure, the total pressure difference across the channel, and the geometric fluidic resistance. By accounting for these considerations, plug coalescence and plug -plug contamination can be eliminated from the splitting process and the symmetry of splitting can be preserved. Furthermore, single-outlet splitting devices were implemented with both valve-and volume-based methods for coordinating the release of output arrays. Arrays of plugs containing commercial sparse matrix screens were obtained from the presented splitting method and these arrays were used in protein crystallization trials. The techniques presented in this paper may facilitate the implementation of high-throughput chemical and biological screening.", "doi": "10.1039/b604993a", "pmcid": "PMC1851925", "issn": "1473-0197", "publisher": "Royal Society of Chemistry", "publication": "Lab on a Chip", "publication_date": "2006", "series_number": "9", "volume": "6", "issue": "9", "pages": "1178-1186" }, { "id": "authors:dgg5g-bk766", "collection": "authors", "collection_id": "dgg5g-bk766", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160735377", "type": "article", "title": "In situ data collection and structure refinement from microcapillary protein crystallization", "author": [ { "family_name": "Yadav", "given_name": "Maneesh K.", "clpid": "Yadav-Maneesh-K" }, { "family_name": "Gerdts", "given_name": "Cory J.", "clpid": "Gerdts-Cory-J" }, { "family_name": "Sanishvili", "given_name": "Ruslan", "clpid": "Sanishvili-Ruslan" }, { "family_name": "Smith", "given_name": "Ward W.", "clpid": "Smith-Ward-W" }, { "family_name": "Roach", "given_name": "L. Spencer", "clpid": "Roach-L-Spencer" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Kuhn", "given_name": "Peter", "clpid": "Kuhn-Peter" }, { "family_name": "Stevens", "given_name": "Raymond C.", "orcid": "0000-0002-4522-8725", "clpid": "Stevens-Raymond-C" } ], "abstract": "In situ X-ray data collection has the potential to eliminate the challenging task of mounting and cryocooling often fragile protein crystals, reducing a major bottleneck in the structure determination process. An apparatus used to grow protein crystals in capillaries and to compare the background X-ray scattering of the components, including thin-walled glass capillaries against Teflon, and various fluorocarbon oils against each other, is described. Using thaumatin as a test case at 1.8 angstrom resolution, this study demonstrates that high-resolution electron density maps and refined models can be obtained from in situ diffraction of crystals grown in microcapillaries.", "doi": "10.1107/S002188980502649X", "pmcid": "PMC1858637", "issn": "0021-8898", "publisher": "International Union of Crystallography", "publication": "Journal of Applied Crystallography", "publication_date": "2005-12", "series_number": "6", "volume": "38", "issue": "6", "pages": "900-905" }, { "id": "authors:2yhp4-m8e50", "collection": "authors", "collection_id": "2yhp4-m8e50", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160727982", "type": "article", "title": "Crystallographic characterization of the geometry changes upon electron loss from 2-tert-butyl-3-aryl-2,3-diazabicyclo 2.2.2 octanes", "author": [ { "family_name": "Nelsen", "given_name": "Stephen F.", "clpid": "Nelsen-S-F" }, { "family_name": "Konradsson", "given_name": "Asgeir E.", "clpid": "Konradsson-A-E" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Guzei", "given_name": "Ilia A.", "clpid": "Guzei-I-A" } ], "abstract": "Crystal structures of 2-tert-butyl-3-(2,3,5,6-tetramethylphenyl)-2,3-diazabicyclo[2.2.2]-octane radical cation nitrate (HyDU+NO_3-) [Hy = (2-tert-butyl-2,3-diazabicyclo[2.2.2]oct-3-yl]; 2-tert-butyl-3-(1-naphthyl)-2,3-diazabicyclo[2.2.2]octane radical cation hexafluoroantiminate (Hy^1NA+SbF_6-); 2-tert-butyl-3-(2-naphthyl)-2,3-diazabicyclo-[2.2.2]octane radical cation hexafluoroantiminate (Hy^2NA+SbF_6-); 1,5-bis(2-tert-butyl-2,3-diazabicyclo[2.2.2]oct-3-yl)naphthalene dication bis(tetraphenylborate) (Hy_2^(15)NA^(2+)(Ph_4B^-)_2); and 2,7-bis(2-tert-butyl-2,3-diazabicyclo[2.2.2]oct-3-yl)naphthalene dication bis(hexafluoroantiminate) (Hy_2^(27)NA^(2+)(SbF_6^-)_2\u00b7CH_3CN) are reported, and the geometries about the oxidized Hy units compared with literature data for neutral Hy-substituted analogues and the geometry changes upon electron loss for these compounds, which have a lone pair, lone pair twist angle in the neutral form (\u03b8(0)) in the range 122\u2212130\u00b0, are compared with those for tetraalkylhydrazines that have \u03b8(0) values near 180, 90, and 0\u00b0.", "doi": "10.1021/cg050154w", "issn": "1528-7483", "publisher": "American Chemical Society", "publication": "Crystal Growth and Design", "publication_date": "2005-11-02", "series_number": "6", "volume": "5", "issue": "6", "pages": "2344-2347" }, { "id": "authors:emdmv-e3886", "collection": "authors", "collection_id": "emdmv-e3886", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160735572", "type": "article", "title": "Using nanoliter plugs in microfluidics to facilitate and understand protein crystallization", "author": [ { "family_name": "Zheng", "given_name": "Bo", "orcid": "0000-0001-8344-3445", "clpid": "Zheng-Bo" }, { "family_name": "Gerdts", "given_name": "Cory J.", "clpid": "Gerdts-Cory-J" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Protein crystallization is important for determining protein structures by X-ray diffraction. Nanoliter-sized plugs aqueous droplets surrounded by a fluorinated carrier fluid have been applied to the screening of protein crystallization conditions. Preformed arrays of plugs in capillary cartridges enable sparse matrix screening. Crystals grown in plugs inside a microcapillary may be analyzed by in situ X-ray diffraction. Screening using plugs, which are easily formed in PDMS microfluidic channels, is simple and economical, and minimizes consumption of the protein. This approach also has the potential to improve our understanding of the fundamentals of protein crystallization, such as the effect of mixing on the nucleation of crystals.", "doi": "10.1016/j.sbi.2005.08.009", "pmcid": "PMC1764865", "issn": "0959-440X", "publisher": "Elsevier", "publication": "Current Opinion in Structural Biology", "publication_date": "2005-10", "series_number": "5", "volume": "15", "issue": "5", "pages": "548-555" }, { "id": "authors:2025f-b6f45", "collection": "authors", "collection_id": "2025f-b6f45", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160717199", "type": "article", "title": "Using microfluidics to observe the effect of mixing on nucleation of protein crystals", "author": [ { "family_name": "Chen", "given_name": "Delai L.", "clpid": "Chen-Delai-L" }, { "family_name": "Gerdts", "given_name": "Cory J.", "clpid": "Gerdts-Cory-J" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper analyzes the effect of mixing on nucleation of protein crystals. The mixing of protein and precipitant was controlled by changing the flow rate in a plug-based microfluidic system. The nucleation rate inversely depended on the flow rate, and flow rate could be used to control nucleation. For example, at higher supersaturations, precipitation happened at low flow rates while large crystals grew at high flow rates. Mixing at low flow velocities in a winding channel induces nucleation more effectively than mixing in straight channels. A qualitative scaling argument that relies on a number of assumptions is presented to understand the experimental results. In addition to helping fundamental understanding, this result may be used to control nucleation, using rapid chaotic mixing to eliminate formation of precipitates at high supersaturation and using slow chaotic mixing to induce nucleation at lower supersaturation.", "doi": "10.1021/ja052279v", "pmcid": "PMC1766325", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2005-07-13", "series_number": "27", "volume": "127", "issue": "27", "pages": "9672-9673" }, { "id": "authors:zdc3e-x1y89", "collection": "authors", "collection_id": "zdc3e-x1y89", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160733236", "type": "article", "title": "Combining microfluidic networks and peptide arrays for multi-enzyme assays", "author": [ { "family_name": "Su", "given_name": "Jing", "clpid": "Su-Jing" }, { "family_name": "Bringer", "given_name": "Michelle R.", "clpid": "Bringer-M-R" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Mrksich", "given_name": "Milan", "clpid": "Mrksich-M" } ], "abstract": "This paper reports the use of microfluidic networks (\u03bcFNs) to both prepare peptide microarrays and carry out label-free enzyme assays on self-assembled monolayers (SAMs) of alkanethiolates on gold. A poly(dimethylsiloxane) (PDMS) stamp fabricated with microchannels is used to immobilize a linear array of cysteine-terminated peptides onto SAMs presenting maleimide groups. The stamp is then reapplied to the SAM in a perpendicular direction to introduce enzyme solutions so that each solution can interact with an identical linear array of immobilized peptides. The \u03bcFNs enable multiple enzyme\u2212substrate interactions to be simultaneously evaluated at a submicroliter scale, while the use of SAMs enables the use of MALDI mass spectrometry (MS) to analyze the enzyme activities. This paper demonstrates applications of this system for assaying multiple kinases and for profiling the activities of kinases and phosphatases in human K562 cell extracts. The combination of \u03bcFN, SAMs, and MS detection provides a flexible platform for assaying enzyme activities in biological samples.", "doi": "10.1021/ja051371o", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2005-05-25", "series_number": "20", "volume": "127", "issue": "20", "pages": "7280-7281" }, { "id": "authors:m1yr2-aqc46", "collection": "authors", "collection_id": "m1yr2-aqc46", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160725036", "type": "article", "title": "Dynamics of Drosophila embryonic patterning network perturbed in space and time using microfluidics", "author": [ { "family_name": "Lucchetta", "given_name": "Elena M.", "clpid": "Lucchetta-Elena-M" }, { "family_name": "Lee", "given_name": "Ji Hwan", "clpid": "Lee-Ji-Hwan" }, { "family_name": "Fu", "given_name": "Lydia A.", "clpid": "Fu-Lydia-A" }, { "family_name": "Patel", "given_name": "Nipam H.", "orcid": "0000-0003-4328-654X", "clpid": "Patel-Nipam-H" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Biochemical networks are perturbed both by fluctuations in environmental conditions and genetic variation. These perturbations must be compensated for, especially when they occur during embryonic pattern formation. Complex chemical reaction networks displaying spatiotemporal dynamics have been controlled and understood by perturbing their environment in space and time. Here, we apply this approach using microfluidics to investigate the robust network in Drosophila melanogaster that compensates for variation in the Bicoid morphogen gradient. We show that the compensation system can counteract the effects of extremely unnatural environmental conditions-a temperature step-in which the anterior and posterior halves of the embryo are developing at different temperatures and thus at different rates. Embryonic patterning was normal under this condition, suggesting that a simple reciprocal gradient system is not the mechanism of compensation. Time-specific reversals of the temperature step narrowed down the critical period for compensation to between 65 and 100 min after onset of embryonic development. The microfluidic technology used here may prove useful to future studies, as it allows spatial and temporal regulation of embryonic development.", "doi": "10.1038/nature03509", "pmcid": "PMC2656922", "issn": "0028-0836", "publisher": "Nature Publishing Group", "publication": "Nature", "publication_date": "2005-04-28", "series_number": "7037", "volume": "434", "issue": "7037", "pages": "1134-1138" }, { "id": "authors:ebrxv-jyt12", "collection": "authors", "collection_id": "ebrxv-jyt12", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160735736", "type": "article", "title": "A Microfluidic Approach for Screening Submicroliter Volumes against Multiple Reagents by Using Preformed Arrays of Nanoliter Plugs in a Three-Phase Liquid/Liquid/Gas Flow", "author": [ { "family_name": "Zheng", "given_name": "Bo", "orcid": "0000-0001-8344-3445", "clpid": "Zheng-Bo" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Plugging a gap in screening: Arrays of nanoliter-sized plugs of different compositions can be preformed in a three-phase liquid/liquid/gas flow. The arrays can be transported into a microfluidic channel to test against a target (see schematic representation), as demonstrated in protein crystallization and an enzymatic assay.", "doi": "10.1002/anie.200462857", "pmcid": "PMC1766320", "issn": "1433-7851", "publisher": "Wiley", "publication": "Angewandte Chemie International Edition", "publication_date": "2005-04-22", "series_number": "17", "volume": "44", "issue": "17", "pages": "2520-2523" }, { "id": "authors:5ck26-hyq79", "collection": "authors", "collection_id": "5ck26-hyq79", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160729676", "type": "article", "title": "Controlling nonspecific protein adsorption in a plug-based microfluidic system by controlling interfacial chemistry using fluorous-phase surfactants", "author": [ { "family_name": "Roach", "given_name": "L. Spencer", "clpid": "Roach-L-Spencer" }, { "family_name": "Song", "given_name": "Helen", "clpid": "Song-Helen" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Control of surface chemistry and protein adsorption is important for using microfluidic devices for biochemical analysis and high-throughput screening assays. This paper describes the control of protein adsorption at the liquid-liquid interface in a plug-based microfluidic system. The microfluidic system uses multiphase flows of immiscible fluorous and aqueous fluids to form plugs, which are aqueous droplets that are completely surrounded by fluorocarbon oil and do not come into direct contact with the hydrophobic surface of the microchannel. Protein adsorption at the aqueous-fluorous interface was controlled by using surfactants; that were soluble in fluorocarbon oil but insoluble in aqueous solutions. Three perfluorinated alkane surfactants capped with different functional groups were used: a carboxylic acid, an alcohol, and a triethylene glycol group that was synthesized from commercially available materials. Using complementary methods of analysis, adsorption was characterized for several proteins (bovine serum albumin (BSA) and fibrinogen), including enzymes (ribonuclease A (RNase A) and alkaline phosphatase). These complementary methods involved characterizing adsorption in microliter-sized droplets by drop tensiometry and in nanoliter plugs by fluorescence microscopy and kinetic measurements of enzyme catalysis. The oligoethylene glycol-capped surfactant prevented protein adsorption in all cases. Adsorption of proteins to the carboxylic acid-capped surfactant in nanoliter plugs could be described by using the Langmuir model and tensiometry results for microliter drops. The microfluidic system was fabricated using rapid prototyping in poly(dimethylsiloxane) (PDMS). Black PDMS microfluidic devices, fabricated by curing a suspension of charcoal in PDMS, were used to measure the changes in fluorescence intensity more sensitively. This system will be useful for microfluidic bioassays, enzymatic kinetics, and protein crystallization, because it does not require surface modification during fabrication to control surface chemistry and protein adsorption.", "doi": "10.1021/ac049061w", "pmcid": "PMC1941690", "issn": "0003-2700", "publisher": "American Chemical Society", "publication": "Analytical Chemistry", "publication_date": "2005-02-01", "series_number": "3", "volume": "77", "issue": "3", "pages": "785-796" }, { "id": "authors:rqf8w-39r07", "collection": "authors", "collection_id": "rqf8w-39r07", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160736036", "type": "article", "title": "Formation of arrayed droplets of soft lithography and two-phase fluid flow, and application in protein crystallization", "author": [ { "family_name": "Zheng", "given_name": "Bo", "orcid": "0000-0001-8344-3445", "clpid": "Zheng-Bo" }, { "family_name": "Tice", "given_name": "Joshua D.", "clpid": "Tice-Joshua-D" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper presents an overview of our recent work on the use of soft lithography and two-phase fluid flow to form arrays of droplets. The crucial issues in the formation of stable arrays of droplets and alternating droplets of two sets of aqueous solutions include the geometry of the microchannels, the capillary number, and the water fraction of the system. Glass capillaries could be coupled to the PDMS microchannels and droplets could be transferred into glass capillaries for long-term storage. The arrays of droplets have been applied to screen the conditions for protein crystallization with microbatch and vapor diffusion techniques.", "doi": "10.1002/adma.200400590", "pmcid": "PMC1858636", "issn": "0935-9648", "publisher": "Wiley", "publication": "Advanced Materials", "publication_date": "2004-08-03", "series_number": "15", "volume": "16", "issue": "15", "pages": "1365-1368" }, { "id": "authors:750pr-xej70", "collection": "authors", "collection_id": "750pr-xej70", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160718167", "type": "article", "title": "A Synthetic Reaction Network: Chemical Amplification Using Nonequilibrium Autocatalytic Reactions Coupled in Time", "author": [ { "family_name": "Gerdts", "given_name": "Cory J.", "clpid": "Gerdts-C-J" }, { "family_name": "Sharoyan", "given_name": "David E.", "clpid": "Sharoyan-D-E" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This article reports a functional chemical reaction network synthesized in a microfluidic device.\nThis chemical network performs chemical 5000-fold amplification and shows a threshold response. It\noperates in a feedforward manner in two stages: the output of the first stage becomes the input of the\nsecond stage. Each stage of amplification is performed by a reaction autocatalytic in Co^(2+). The microfluidic\nnetwork is used to maintain the two chemical reactions away from equilibrium and control the interactions\nbetween them in time. Time control is achieved as described previously (Angew. Chem., Int. Ed. 2003, 42,\n768) by compartmentalizing the reaction mixture inside plugs which are aqueous droplets carried through\na microchannel by an immiscible fluorinated fluid. Autocatalytic reaction displayed sensitivity to mixing;\nmore rapid mixing corresponded to slower reaction rates. Synthetic chemical reaction networks may help\nunderstand the function of biochemical reaction networks, the goal of systems biology. They may also find\npractical applications. For example, the system described here may be used to detect visually, in a simple\nformat, picoliter volumes of nanomolar concentrations of Co^(2+), an environmental pollutant.", "doi": "10.1021/ja031689l", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2004-05-26", "series_number": "20", "volume": "126", "issue": "20", "pages": "6327-6331" }, { "id": "authors:rsr0p-kcs98", "collection": "authors", "collection_id": "rsr0p-kcs98", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160716511", "type": "article", "title": "Microfluidic Systems for Chemical Kinetics that Rely on Chaotic Mixing in Droplets", "author": [ { "family_name": "Bringer", "given_name": "Michelle R.", "clpid": "Bringer-Michelle-R" }, { "family_name": "Gerdts", "given_name": "Cory J.", "clpid": "Gerdts-Cory-J" }, { "family_name": "Song", "given_name": "Helen", "clpid": "Song-Helen" }, { "family_name": "Tice", "given_name": "Joshua D.", "clpid": "Tice-Joshua-D" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper reviews work on a microfluidic system that relies on chaotic advection\nto rapidly mix multiple reagents isolated in droplets (plugs). Using a combination\nof turns and straight sections, winding microfluidic channels create unsteady fluid\nflows that rapidly mix the multiple reagents contained within plugs. The scaling\nof mixing for a range of channel widths, flow velocities and diffusion coefficients\nhas been investigated. Due to rapid mixing, low sample consumption and transport\nof reagents with no dispersion, the system is particularly appropriate for chemical\nkinetics and biochemical assays. The mixing occurs by chaotic advection and is rapid\n(sub-millisecond), allowing for an accurate description of fast reaction kinetics. In\naddition, mixing has been characterized and explicitly incorporated into the kinetic\nmodel.", "doi": "10.1098/rsta.2003.1364", "pmcid": "PMC1769314", "issn": "1364-503X", "publisher": "Royal Society of London", "publication": "Philosophical Transactions A: Mathematical, Physical and Engineering Sciences", "publication_date": "2004-05-15", "series_number": "1818", "volume": "362", "issue": "1818", "pages": "1087-1104" }, { "id": "authors:bhw79-ptr61", "collection": "authors", "collection_id": "bhw79-ptr61", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160736358", "type": "article", "title": "A Droplet-Based, Composite PDMS/Glass Capillary Microfluidic System for Evaluating Protein Crystallization Conditions by Microbatch and Vapor-Diffusion Methods with On-Chip X-Ray Diffraction", "author": [ { "family_name": "Zheng", "given_name": "Bo", "orcid": "0000-0001-8344-3445", "clpid": "Zheng-Bo" }, { "family_name": "Tice", "given_name": "Joshua D.", "clpid": "Tice-Joshua-D" }, { "family_name": "Roach", "given_name": "L. Spencer", "clpid": "Roach-L-Spencer" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "On-chip protein crystallization and diffraction: Proteins can be crystallized in nanoliter volumes by using both microbatch and vapor-diffusion techniques inside composite PDMS/glass microfluidic devices (PDMS=polydimethylsiloxane). The quality of crystals can be assessed directly by on-chip X-ray diffraction.", "doi": "10.1002/anie.200453974", "pmcid": "PMC1766324", "issn": "1433-7851", "publisher": "Wiley", "publication": "Angewandte Chemie International Edition", "publication_date": "2004-05-03", "series_number": "19", "volume": "43", "issue": "19", "pages": "2508-2511" }, { "id": "authors:qvpvf-4wf12", "collection": "authors", "collection_id": "qvpvf-4wf12", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160733899", "type": "article", "title": "Effects of viscosity on droplet formation and mixing in microfluidic channels", "author": [ { "family_name": "Tice", "given_name": "J. D.", "clpid": "Tice-J-D" }, { "family_name": "Lyon", "given_name": "A. D.", "clpid": "Lyon-A-D" }, { "family_name": "Ismagilov", "given_name": "R. F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper characterizes the conditions required to form nanoliter-sized droplets (plugs) of viscous aqueous reagents in flows of inuniscible carrier fluid within microfluidic channels. For both non-viscous (viscosity of 2.0 mPa s) and viscous (viscosity of 18 mPa s) aqueous solutions, plugs formed reliably in a flow of water-immiscible carrier fluid for Capillary number less than 0.01, although plugs were able to form at higher Capillary numbers at lower ratios of the aqueous phase flow rate to the flow rate of the carrier fluid (in all the experiments performed, the Reynolds number was less than 1). The paper also shows that combining viscous and non-viscous reagents can enhance mixing in droplets moving through straight microchannels by providing a nearly ideal initial distribution of reagents within each droplet. The study should facilitate the use of this droplet-based microfluidic platform for investigation of protein crystallization, kinetics, and assays.", "doi": "10.1016/j.aca.2003.11.024", "issn": "0003-2670", "publisher": "Elsevier", "publication": "Analytica Chimica Acta", "publication_date": "2004-04-01", "series_number": "1", "volume": "507", "issue": "1", "pages": "73-77" }, { "id": "authors:gw0cd-hyp12", "collection": "authors", "collection_id": "gw0cd-hyp12", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160729811", "type": "article", "title": "Minimal Functional Model of Hemostasis in a Biomimetic Microfluidic System", "author": [ { "family_name": "Runyon", "given_name": "Matthew K.", "clpid": "Runyon-M-K" }, { "family_name": "Johnson-Kerner", "given_name": "Bethany L.", "clpid": "Johnson-Kerner-B-L" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "The proof of the model is in the function: A minimal model of hemostasis (a complex biochemical network responsible for blood coagulation) may be implemented with only three chemical reactions, which creates a biomimetic functional microfluidic system that is capable of repairing itself (as modeled in the figure). This simple system shows threshold response and sensitivity to flow similar to that observed in hemostasis.", "doi": "10.1002/anie.200353428", "issn": "1433-7851", "publisher": "Wiley", "publication": "Angewandte Chemie International Edition", "publication_date": "2004-03-12", "series_number": "12", "volume": "43", "issue": "12", "pages": "1531-1536" }, { "id": "authors:jwp5s-20346", "collection": "authors", "collection_id": "jwp5s-20346", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160731532", "type": "article", "title": "Multi-step synthesis of nanoparticles performed on millisecond time scale in a microfluidic droplet-based system", "author": [ { "family_name": "Shestopalov", "given_name": "Ilya", "clpid": "Shestopalov-I" }, { "family_name": "Tice", "given_name": "Joshua D.", "clpid": "Tice-J-D" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper reports a plug-based, microfluidic method for performing multi-step chemical reactions with millisecond\ntime-control. It builds upon a previously reported method where aqueous reagents were injected into a flow of\nimmiscible fluid (fluorocarbons) (H. Song et al., Angew. Chem. Int. Ed., 2003, 42, 768). The aqueous reagents formed\nplugs \u2013 droplets surrounded and transported by the immiscible fluid. Winding channels rapidly mixed the reagents in\ndroplets. This paper shows that further stages of the reaction could be initiated by flowing additional reagent streams\ndirectly into the droplets of initial reaction mixture. The conditions necessary for an aqueous stream to merge with\naqueous droplets were characterized. The Capillary number could be used to predict the behavior of the two-phase flow\nat the merging junction. By transporting solid reaction products in droplets, the products were kept from aggregating on\nthe walls of the microchannels. To demonstrate the utility of this microfluidic method it was used to synthesize colloidal\nCdS and CdS/CdSe core-shell nanoparticles.", "doi": "10.1039/B403378G", "issn": "1473-0197", "publisher": "Royal Society of Chemistry", "publication": "Lab on a Chip", "publication_date": "2004", "series_number": "4", "volume": "4", "issue": "4", "pages": "316-321" }, { "id": "authors:t0afb-m2818", "collection": "authors", "collection_id": "t0afb-m2818", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160736217", "type": "article", "title": "Formation of droplets of alternating composition in microfluidic channels and applications to indexing of concentrations in droplet-based assays", "author": [ { "family_name": "Zheng", "given_name": "Bo", "orcid": "0000-0001-8344-3445", "clpid": "Zheng-Bo" }, { "family_name": "Tice", "given_name": "Joshua D.", "clpid": "Tice-Joshua-D" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "For screening the conditions for a reaction by using droplets (or plugs) as microreactors, the composition of the droplets must be indexed. Indexing here refers to measuring the concentration of a solute by addition of a marker, either internal or external. Indexing may be performed by forming droplet pairs, where in each pair the first droplet is used to conduct the reaction, and the second droplet is used to index the composition of the first droplet. This paper characterizes a method for creating droplet pairs by generating alternating droplets, of two sets of aqueous solutions in a flow of immiscible carrier fluid within PDMS and glass microfluidic channels. The paper also demonstrates that the technique can be used to index the composition of the droplets, and this application is illustrated by screening conditions of protein crystallization. The fluid properties required to form the steady flow of the alternating droplets in a microchannel were characterized as a function of the capillary number Ca and water fraction. Four regimes were observed. At the lowest values of Ca, the droplets of the two streams coalesced; at intermediate values of Ca the alternating droplets formed reliably. At even higher values of Ca, shear forces dominated and caused formation of droplets that were smaller than the cross-sectional dimension of the channel; at the highest values of Ca, coflowing laminar streams of the two immiscible fluids formed. In addition to screening of protein crystallization conditions, understanding of the fluid flow in this system may extend this indexing approach to other chemical and biological assays performed on a microfluidic chip.", "doi": "10.1021/ac0495743", "pmcid": "PMC1766978", "issn": "0003-2700", "publisher": "American Chemical Society", "publication": "Analytical Chemistry", "publication_date": "2004", "series_number": "17", "volume": "76", "issue": "17", "pages": "4977-4982" }, { "id": "authors:exqg6-0r797", "collection": "authors", "collection_id": "exqg6-0r797", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160732315", "type": "article", "title": "Millisecond kinetics on a microfluidic chip using nanoliters of reagents", "author": [ { "family_name": "Song", "given_name": "Helen", "clpid": "Song-Helen" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper describes a microfluidic chip for performing kinetic measurements with better than millisecond resolution. Rapid kinetic measurements in microfluidic systems are complicated by two problems: mixing is slow and dispersion is large. These problems also complicate biochemical assays performed in microfluidic chips. We have recently shown (Song, H.; Tice, J. D.; Ismagilov, R. F. Angew. Chem., Int. Ed. 2003, 42, 768-772) how multiphase fluid flow in microchannels can be used to address both problems by transporting the reagents inside aqueous droplets (plugs) surrounded by an immiscible fluid. Here, this droplet-based microfluidic system was used to extract kinetic parameters of an enzymatic reaction. Rapid single-turnover kinetics of ribonuclease A (RNase A) was measured with better than millisecond resolution using sub-microliter volumes of solutions. To obtain the single-turnover rate constant (k = 1100 +/- 250 s(-1)), four new features for this microfluidics platform were demonstrated: (i) rapid on-chip dilution, (ii) multiple time range access, (iii) biocompatibility with RNase A, and (iv) explicit treatment of mixing for improving time resolution of the system. These features are discussed using kinetics of RNase A. From fluorescent images integrated for 2-4 s, each kinetic profile can be obtained using less than 150 nL of solutions of reagents because this system relies on chaotic advection inside moving droplets rather than on turbulence to achieve rapid mixing. Fabrication of these devices in PDMS is straightforward and no specialized equipment, except for a standard microscope with a CCD camera, is needed to run the experiments. This microfluidic platform could serve as an inexpensive and economical complement to stopped-flow methods for a broad range of time-resolved experiments and assays in chemistry and biochemistry.", "doi": "10.1021/ja0354566", "pmcid": "PMC1769313", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2003-11-26", "series_number": "47", "volume": "125", "issue": "47", "pages": "14613-14619" }, { "id": "authors:012af-kmz12", "collection": "authors", "collection_id": "012af-kmz12", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160734086", "type": "article", "title": "Formation of droplets and mixing in multiphase microfluidics at low values of the Reynolds and the capillary numbers", "author": [ { "family_name": "Tice", "given_name": "Joshua D.", "clpid": "Tice-J-D" }, { "family_name": "Song", "given_name": "Helen", "clpid": "Song-Helen" }, { "family_name": "Lyon", "given_name": "Adam D.", "clpid": "Lyon-A-D" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This paper reports an experimental characterization of a simple method for rapid formation of droplets, or plugs, of multiple aqueous reagents without bringing reagents into contact prior to mixing. Droplet-based microfluidics offers a simple method of achieving rapid mixing and transport with no dispersion. In addition, this paper shows that organic dyes at high concentrations should not be used for the visualization of flow patterns and mixing of aqueous plugs in multiphase flows in this system (fluorinated carrier fluid and PDMS microchannels). It reports an inorganic dye that can be used instead. This work focuses on mixing in plugs moving through straight channels. It demonstrates that, when traveling through straight microchannels, mixing within plugs by steady recirculating flow is highly sensitive to the initial distribution of the aqueous reagents established by the eddy flow at the tip of the forming plug (twirling). The results also show how plugs with proper distribution of the aqueous reagents could be formed in order to achieve optimal mixing of the reagents in this system.", "doi": "10.1021/la030090w", "issn": "0743-7463", "publisher": "American Chemical Society", "publication": "Langmuir", "publication_date": "2003-10-28", "series_number": "22", "volume": "19", "issue": "22", "pages": "9127-9133" }, { "id": "authors:vt9r8-5sp43", "collection": "authors", "collection_id": "vt9r8-5sp43", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160735877", "type": "article", "title": "Screening of protein crystallization conditions on a microfluidic chip using nanoliter-size droplets", "author": [ { "family_name": "Zheng", "given_name": "Bo", "orcid": "0000-0001-8344-3445", "clpid": "Zheng-Bo" }, { "family_name": "Roach", "given_name": "L. Spencer", "clpid": "Roach-L-S" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Protein crystallization is a major bottleneck in determining tertiary protein structures from genomic sequence data. This paper describes a microfluidic system for screening hundreds of protein crystallization conditions using less than 4 nL of protein solution for each crystallization droplet. The droplets are formed by mixing protein, precipitant, and additive stock solutions in variable ratios in a flow of water-immiscible fluids inside microchannels. Each droplet represents a discrete trial testing different conditions. The system has been validated by crystallization of several water-soluble proteins.", "doi": "10.1021/ja037166v", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2003-09-17", "series_number": "37", "volume": "125", "issue": "37", "pages": "11170-11171" }, { "id": "authors:g73kc-wpk51", "collection": "authors", "collection_id": "g73kc-wpk51", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160719583", "type": "article", "title": "Integrated microfluidic systems", "author": [ { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Microchips: Multistep reactions may be catalyzed by enzymes immobilized on microbeads trapped inside microfluidic channels. Large integrated microfluidic circuits can be created by using multilayer soft lithography in poly(dimethylsiloxane). These circuits (see schematic representation) can be used to perform hundreds of different reactions simultaneously, and can be applied to a range of problems, from enzymatic assays to crystallization of proteins.", "doi": "10.1002/anie.200301660", "issn": "1433-7851", "publisher": "Wiley", "publication": "Angewandte Chemie International Edition", "publication_date": "2003-09-15", "series_number": "35", "volume": "42", "issue": "35", "pages": "4130-4132" }, { "id": "authors:6gnkw-hmq18", "collection": "authors", "collection_id": "6gnkw-hmq18", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160732896", "type": "article", "title": "Fluidic Ratchet Based on Marangoni\u2212B\u00e9nard Convection", "author": [ { "family_name": "Stroock", "given_name": "Abraham D.", "clpid": "Stroock-A-D" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Stone", "given_name": "Howard A.", "clpid": "Stone-H-A" }, { "family_name": "Whitesides", "given_name": "George M.", "orcid": "0000-0001-9451-2442", "clpid": "Whitesides-G-M" } ], "abstract": "A mean flow is observed experimentally in a layer of fluid undergoing Marangoni-Benard convection over a heated substrate that presents a pattern of asymmetrical grooves. The direction of the mean flow is a function of the temperature difference across the layer and of the thickness of the layer; the direction can be controlled by changing these parameters. This system acts as a fluidic ratchet: the local structure of the thermally driven convection interacts with the asymmetry of the local topographical pattern and causes a net, global flow in the fluid. This fluidic ratchet may be useful in handling fluids on macroscopic and microscopic scales.", "doi": "10.1021/la026400c", "issn": "0743-7463", "publisher": "American Chemical Society", "publication": "Langmuir", "publication_date": "2003-05-13", "series_number": "10", "volume": "19", "issue": "10", "pages": "4358-4362" }, { "id": "authors:b1nsr-ck282", "collection": "authors", "collection_id": "b1nsr-ck282", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160732735", "type": "article", "title": "A Microfluidic System for Controlling Reaction Networks in Time", "author": [ { "family_name": "Song", "given_name": "Helen", "clpid": "Song-Helen" }, { "family_name": "Tice", "given_name": "Joshua D.", "clpid": "Tice-J-D" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Millisecond mixing and transport with no dispersion are achieved by unsteady flows induced in droplets of about 60 pL that travel through winding microfluidic channels (top). Fluorescence can be used to monitor mixing (bottom) or measure reaction rates. In principle, arbitrarily complex reaction networks can be created by combining and splitting streams of such droplets.", "doi": "10.1002/anie.200390203", "issn": "1433-7851", "publisher": "Wiley", "publication": "Angewandte Chemie International Edition", "publication_date": "2003-02-17", "series_number": "7", "volume": "42", "issue": "7", "pages": "768-772" }, { "id": "authors:7m9bq-gfk14", "collection": "authors", "collection_id": "7m9bq-gfk14", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160731877", "type": "article", "title": "Experimental test of scaling of mixing by chaotic advection in droplets moving through microfluidic channels", "author": [ { "family_name": "Song", "given_name": "Helen", "clpid": "Song-Helen" }, { "family_name": "Bringer", "given_name": "Michelle R.", "clpid": "Bringer-Michelle-R" }, { "family_name": "Tice", "given_name": "Joshua D.", "clpid": "Tice-Joshua-D" }, { "family_name": "Gerdts", "given_name": "Cory J.", "clpid": "Gerdts-Cory-J" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "This letter describes an experimental test of a simple argument that predicts the scaling of chaotic mixing in a droplet moving through a winding microfluidic channel. Previously, scaling arguments for chaotic mixing have been described for a flow that reduces striation length by stretching, folding, and reorienting the fluid in a manner similar to that of the baker's transformation. The experimentally observed flow patterns within droplets (or plugs) resembled the baker's transformation. Therefore, the ideas described in the literature could be applied to mixing in droplets to obtain the scaling argument for the dependence of the mixing time, t \u223c (aw/U)log(Pe), where w [m] is the cross-sectional dimension of the microchannel, a is the dimensionless length of the plug measured relative to w, U [m\u200as^\u22121] is the flow velocity, Pe is the P\u00e9clet number (Pe = wU/D), and D [m^2\u200as^\u22121] is the diffusion coefficient of the reagent being mixed. Experiments were performed to confirm the scaling argument by varying the parameters w, U, and D. Under favorable conditions, submillisecond mixing has been demonstrated in this system.", "doi": "10.1063/1.1630378", "pmcid": "PMC2025702", "issn": "0003-6951", "publisher": "American Institute of Physics", "publication": "Applied Physics Letters", "publication_date": "2003", "series_number": "22", "volume": "83", "issue": "22", "pages": "4664-4666" }, { "id": "authors:hpe9w-e2462", "collection": "authors", "collection_id": "hpe9w-e2462", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160720419", "type": "article", "title": "Autonomous movement and self-assembly", "author": [ { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Schwartz", "given_name": "Alexander", "clpid": "Schwartz-A" }, { "family_name": "Bowden", "given_name": "Ned", "clpid": "Bowden-N" }, { "family_name": "Whitesides", "given_name": "George M.", "orcid": "0000-0001-9451-2442", "clpid": "Whitesides-G-M" } ], "abstract": "The artificial millimeter-scale \"autonomous movers\" glide across the surface of a liquid without an external power source. This system is based on a combination of two processes: Motion of individual objects powered by the catalytic decomposition of hydrogen peroxide, and relative motion (self-assembly) caused by capillary interactions at the fluid/air interface. The picture shows the rotational/translational motion of a single object; the motion of a pair of these object depends on their chirality.", "doi": "10.1002/1521-3773(20020215)41:4<652::AID-ANIE652>3.0.CO;2-U", "issn": "1433-7851", "publisher": "Wiley", "publication": "Angewandte Chemie International Edition", "publication_date": "2002-02-15", "series_number": "4", "volume": "41", "issue": "4", "pages": "652-654" }, { "id": "authors:fmpcf-bej39", "collection": "authors", "collection_id": "fmpcf-bej39", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160719890", "type": "article", "title": "Microfluidic arrays of fluid-fluid diffusional contacts as detection elements and combinatorial tools", "author": [ { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Ng", "given_name": "Jessamine M. K.", "clpid": "Ng-Jessamine-M-K" }, { "family_name": "Kenis", "given_name": "Paul J. A.", "clpid": "Kenis-P-J-A" }, { "family_name": "Whitesides", "given_name": "George M.", "orcid": "0000-0001-9451-2442", "clpid": "Whitesides-G-M" } ], "abstract": "This paper describes microfluidic systems that can be used to investigate multiple chemical or biochemical interactions in a parallel format. These three-dimensional systems are generated by crossing two sets of microfluidic channels, fabricated in two different layers, at tight angles. Solutions of the reagents are placed in the channels; in different modes of operation, these solutions can be either flowing or stationary- the latter is important when one set of channels is filled with viscous gels with immobilized reagents. At every crossing, the channels are separated either by a single membrane or by a composite separator comprising a membrane, a microwell, and a second membrane. These components allow diffusive mass transport and minimize convective transport through the crossing. Polycarbonate membranes with 0.1-1-mum vertical pores were used to fabricate the devices. Each crossing of parallel channels serves as an element in which chemical or biochemical interactions can take place; interactions can be detected by monitoring changes in fluorescence and absorbance. These all- organic systems are straightforward to fabricate and to operate and may find applications as portable microanalytical systems and as tools in combinatorial research.", "doi": "10.1021/ac010502a", "issn": "0003-2700", "publisher": "American Chemical Society", "publication": "Analytical Chemistry", "publication_date": "2001-11-01", "series_number": "21", "volume": "73", "issue": "21", "pages": "5207-5213" }, { "id": "authors:warvr-y6x22", "collection": "authors", "collection_id": "warvr-y6x22", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160720235", "type": "article", "title": "Pressure-driven laminar flow in tangential microchannels: an elastomeric microfluidic switch", "author": [ { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Rosmarin", "given_name": "David", "clpid": "Rosmarin-D" }, { "family_name": "Kenis", "given_name": "Paul J. A.", "clpid": "Kenis-P-J-A" }, { "family_name": "Chiu", "given_name": "Daniel T.", "clpid": "Chiu-Daniel-T" }, { "family_name": "Zhang", "given_name": "Wendy", "clpid": "Zhang-Wendy" }, { "family_name": "Stone", "given_name": "Howard A.", "clpid": "Stone-H-A" }, { "family_name": "Whitesides", "given_name": "George M.", "orcid": "0000-0001-9451-2442", "clpid": "Whitesides-G-M" } ], "abstract": "This paper describes laminar fluid flow through a three- dimensional elastomeric microstructure formed by two microfluidic channels, fabricated in layers that contact one another face-to-face (typically at a 90 degrees angle), with the fluid flows in tangential contact. There are two ways to control fluid flow through these tangentially connected microchannels. First, the flow profiles through the crossings are sensitive to the aspect ratio of the channels; the flow can be controlled by applying external pressure and changing this aspect ratio. Second, the flow direction of an individual laminar stream in multiphase laminar flow depends on the lateral position of the stream within the channel; this position can be controlled by injecting additional streams of fluid into the channel. We describe two microfluidic switches based on these two ways for controlling fluid flow through tangential microchannels and present theoretical arguments that explain the observed dependence of the flow profiles on the aspect ratio of the channels.", "doi": "10.1021/ac010374q", "issn": "0003-2700", "publisher": "American Chemical Society", "publication": "Analytical Chemistry", "publication_date": "2001-10-01", "series_number": "19", "volume": "73", "issue": "19", "pages": "4682-4687" }, { "id": "authors:2vtmg-1y153", "collection": "authors", "collection_id": "2vtmg-1y153", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160720076", "type": "article", "title": "Competition of intrinsic and topographically imposed patterns in Benard-Marangoni convection", "author": [ { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Rosmarin", "given_name": "David", "clpid": "Rosmarin-D" }, { "family_name": "Gracias", "given_name": "David H.", "clpid": "Gracias-D-H" }, { "family_name": "Stroock", "given_name": "Abraham D.", "clpid": "Stroock-A-D" }, { "family_name": "Whitesides", "given_name": "George M.", "orcid": "0000-0001-9451-2442", "clpid": "Whitesides-G-M" } ], "abstract": "The structure of Benard-Marangoni convection cells can be controlled by periodic topographic patterns on the heated surface that generates the convection. When the periodicity of the topographic pattern matches the intrinsic periodicity of the convection cells, a convective pattern is formed that is 1:1 commensurate with the topographic pattern. Arrays of hexagonal, square, and triangular convection cells were generated over the appropriately designed topographic patterns, and visualized by infrared imaging. For imposed patterns with periodicity in two dimensions, as the ratio of the intrinsic and perturbing length scales changes, the pattern of the convection cells shows sharp transitions between different patterns commensurate with the imposed pattern. For imposed patterns with periodicity in one dimension (i.e., lines) the convection cells use the unconstrained dimension to adapt continuously to the external perturbation. Topographically controlled convection cells can be used to assemble microscopic particles into externally switchable regular lattices. (C) 2001 American Institute of Physics.", "doi": "10.1063/1.1384473", "issn": "0003-6951", "publisher": "American Institute of Physics", "publication": "Applied Physics Letters", "publication_date": "2001-07-16", "series_number": "3", "volume": "79", "issue": "3", "pages": "439-441" }, { "id": "authors:pp34s-cre17", "collection": "authors", "collection_id": "pp34s-cre17", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160728530", "type": "article", "title": "Solvent effects on charge transfer bands of nitrogen-centered intervalence compounds", "author": [ { "family_name": "Nelsen", "given_name": "Stephen F.", "clpid": "Nelsen-S-F" }, { "family_name": "Trieber", "given_name": "Dwight A., II", "clpid": "Trieber-D-A-II" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Teki", "given_name": "Yoshio", "clpid": "Teki-Y" } ], "abstract": "Electron transfer parameters are extracted from the optical spectra of intervalence bis(hydrazine) radical cations. Compounds with 2-tert-butyl-3-phenyl-2,3-diazabicyclo[2.2.2]octyl-containing charge-bearing units that are doubly linked by 4-\u03c3-bond and by 6-\u03c3-bond saturated bridges are compared with ones having tert-butylisopropyl- and diphenyl-substituted charge bearing units and others having the aromatic units functioning as the bridge. Solvent effect studies show that the optical transition energy (E_(op)) does not behave as dielectric continuum theory predicts but that solvent reorganization energy may be usefully separated from the vibrational reorganization energy by including linear terms in both the Pekar factor (\u03b3) and the Gutmann donor number (DN) in correlating the solvent effect. Solvation of the bridge for these compounds is too large to ignore, which makes dielectric continuum theory fail to properly predict solvent effects on either E_(op) or the free energy for comproportionation.", "doi": "10.1021/ja003436n", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2001-06-20", "series_number": "24", "volume": "123", "issue": "24", "pages": "5684-5694" }, { "id": "authors:b1ede-7p147", "collection": "authors", "collection_id": "b1ede-7p147", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160718857", "type": "article", "title": "Correlating electron transport and molecular structure in organic thin films", "author": [ { "family_name": "Holmlin", "given_name": "R. Erik", "clpid": "Holmlin-R-E" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Haag", "given_name": "Rainer", "clpid": "Haag-R" }, { "family_name": "Mujica", "given_name": "Vladimiro", "clpid": "Mujica-V" }, { "family_name": "Ratner", "given_name": "Mark A.", "clpid": "Ratner-M-A" }, { "family_name": "Rampi", "given_name": "Maria Anita", "clpid": "Rampi-M-A" }, { "family_name": "Whitesides", "given_name": "George M.", "orcid": "0000-0001-9451-2442", "clpid": "Whitesides-G-M" } ], "abstract": "A convenient experimental system is described, with which electron transport through structurally well-defined, 2\u20135 nm-thick, organic films can be examined. Two types of junction J have been studied in which self-assembled monolayers (SAMs, for example, SAM(1) formed on Ag from aliphatic and aromatic thiols, and SAM(2), formed on Hg from hexadecanethiol) are in contact through either van der Waals interactions (see picture) or through covalent, hydrogen, or ionic bonds.", "doi": "10.1002/1521-3773(20010618)40:12<2316::AID-ANIE2316>3.0.CO;2-#", "issn": "1433-7851", "publisher": "Wiley", "publication": "Angewandte Chemie International Edition", "publication_date": "2001-06-18", "series_number": "12", "volume": "40", "issue": "12", "pages": "2316-2320" }, { "id": "authors:wwktg-f5w34", "collection": "authors", "collection_id": "wwktg-f5w34", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160718693", "type": "article", "title": "Electron transport through thin organic films in metal- insulator-metal junctions based on self-assembled monolayers", "author": [ { "family_name": "Holmlin", "given_name": "R. Erik", "clpid": "Holmlin-R-E" }, { "family_name": "Haag", "given_name": "Rainer", "clpid": "Haag-R" }, { "family_name": "Chabinyc", "given_name": "Michael L.", "orcid": "0000-0003-4641-3508", "clpid": "Chabinyc-M-L" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Cohen", "given_name": "Adam E.", "clpid": "Cohen-A-E" }, { "family_name": "Terfort", "given_name": "Andreas", "clpid": "Terfort-A" }, { "family_name": "Rampi", "given_name": "Maria Anita", "clpid": "Rampi-M-A" }, { "family_name": "Whitesides", "given_name": "George M.", "orcid": "0000-0001-9451-2442", "clpid": "Whitesides-G-M" } ], "abstract": "This paper describes an experimentally simple system for measuring rates of electron transport across organic thin films having a range of molecular structures. The system uses a metal\u2212insulator\u2212metal junction based on self-assembled monolayers (SAMs); it is particularly easy to assemble. The junction consists of a SAM supported on a silver film (Ag-SAM(1)) in contact with a second SAM supported on the surface of a drop of mercury (Hg-SAM(2))that is, a Ag-SAM(1)SAM(2)-Hg junction. SAM(1) and SAM(2) can be derived from the same or different thiols. The current that flowed across junctions with SAMs of aliphatic thiols or aromatic thiols on Ag and a SAM of hexadecane thiol on Hg depended both on the molecular structure and on the thickness of the SAM on Ag:\u2009 the current density at a bias of 0.5 V ranged from 2 \u00d7 10^-10 A/cm^2 for HS(CH_2)_15CH_3 on Ag to 1 \u00d7 10^-6 A/cm^2 for HS(CH_2)_7CH_3 on Ag, and from 3 \u00d7 10^-6 A/cm^2 for HS(Ph)_3H (Ph = 1,4-C_6H4_) on Ag to 7 \u00d7 10^-4 A/cm^2 for HSPhH on Ag. The current density increased roughly linearly with the area of contact between SAM(1) and SAM(2), and it was not different between Ag films that were 100 or 200 nm thick. The current\u2212voltage curves were symmetrical around V = 0. The current density decreased with increasing distance between the electrodes according to the relation I = I0e-\u03b2dAg,Hg, where dAg,Hg is the distance between the electrodes, and \u03b2 is the structure-dependent attenuation factor for the molecules making up SAM(1). At an applied potential of 0.5 V, \u03b2 was 0.87 \u00b1 0.1 \u00c5-1 for alkanethiols, 0.61 \u00b1 0.1 \u00c5-1 for oligophenylene thiols, and 0.67 \u00b1 0.1 \u00c5-1 for benzylic derivatives of oligophenylene thiols. The values of \u03b2 did not depend significantly on applied potential over the range of 0.1 to 1 V. These junctions provide a test bed with which to screen the intrinsic electrical properties of SAMs made up of molecules with different structures; information obtained using these junctions will be useful in correlating molecular structure and rates of electron transport.", "doi": "10.1021/ja004055c", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "2001-05-30", "series_number": "21", "volume": "123", "issue": "21", "pages": "5075-5085" }, { "id": "authors:vkf5k-mvj68", "collection": "authors", "collection_id": "vkf5k-mvj68", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160721858", "type": "article", "title": "Fabrication inside microchannels using fluid flow", "author": [ { "family_name": "Kenis", "given_name": "Paul J. A.", "clpid": "Kenis-P-J-A" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Takayama", "given_name": "Shuichi", "clpid": "Takayama-Shuichi" }, { "family_name": "Whitesides", "given_name": "George M.", "orcid": "0000-0001-9451-2442", "clpid": "Whitesides-G-M" }, { "family_name": "Li", "given_name": "Shulong", "clpid": "Li-Shulong" }, { "family_name": "White", "given_name": "Henry S.", "clpid": "White-H-S" } ], "abstract": "This Account summarizes techniques for carrying out microfabrication of structures with dimensions down to 10 mum in microchannels that are 0.02-2 mm wide. These methods are largely based on the exploitation of laminar flow at low Reynolds number (Re) to control the spatial delivery of reagents. These methods are illustrated by fabrication of fibers, microelectrode arrays, arrays of crystals, and patterns of proteins and cells.", "doi": "10.1021/ar000062u", "issn": "0001-4842", "publisher": "American Chemical Society", "publication": "Accounts of Chemical Research", "publication_date": "2000-12-19", "series_number": "12", "volume": "33", "issue": "12", "pages": "841-847" }, { "id": "authors:ydewe-tsv40", "collection": "authors", "collection_id": "ydewe-tsv40", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160719216", "type": "article", "title": "Evidence for a two-step electron-transfer process in the electrode reactions of tetraisopropylhydrazine, tetracyclohexylhydrazine and their radical cation salts", "author": [ { "family_name": "Hong", "given_name": "Sun Hee", "clpid": "Hong-Sun-Hee" }, { "family_name": "Evans", "given_name": "Dennis H.", "clpid": "Evans-D-H" }, { "family_name": "Nelsen", "given_name": "Stephen F.", "clpid": "Nelsen-S-F" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "The heterogeneous electron-transfer kinetics of tetraisopropylhydrazine (4), tetraisopropylhydrazine radical cation hexafluoroantimonate (4+radical dot SbF_6^\u2212), tetracyclohexylhydrazine (5) and tetracyclohexylhydrazine radical cation hexafluoroantimonate (5+radical dot SbF_6^\u2212) have been studied by cyclic voltammetry at a gold working electrode in acetonitrile containing 0.10 M tetrabutylammonium hexafluorophosphate. Results were obtained at eight scan rates between 0.2 and 40 V s^\u22121 and six temperatures ranging from \u221215 to 50\u00b0C. The results were analyzed according to two models: (1) A direct, one-step electron transfer in which structural change and electron transfer are concerted. (2) A two-step process in which structural change is considered as a separate chemical reaction that precedes or follows the electron-transfer event. Specifically, a square scheme is proposed in which the favored untwisted radical cation can convert to a twisted version, which then receives an electron to form the favored twisted neutral hydrazine. Also, the favored twisted neutral can convert to an untwisted version, which can give up an electron to form the favored untwisted radical cation, thus completing the square. It was found that the one-step model was unable to account for the voltammetric data. On the other hand, analysis by the two-step mechanism produced substantially better agreement between simulation and experiment, particularly for 4 and 4+radical dot SbF_6^\u2212. The present experiments provide the first evidence that two-step electron transfer reactions occur with acyclic tetraalkylhydrazines.", "doi": "10.1016/S0022-0728(00)00137-6", "issn": "0022-0728", "publisher": "Elsevier", "publication": "Journal of Electroanalytical Chemistry", "publication_date": "2000-05-22", "series_number": "1", "volume": "486", "issue": "1", "pages": "75-84" }, { "id": "authors:w6fvd-m9p56", "collection": "authors", "collection_id": "w6fvd-m9p56", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160733045", "type": "article", "title": "Patterning electro-osmotic flow with patterned surface charge", "author": [ { "family_name": "Stroock", "given_name": "Abraham D.", "clpid": "Stroock-A-D" }, { "family_name": "Weck", "given_name": "Marcus", "clpid": "Weck-M" }, { "family_name": "Chiu", "given_name": "Daniel T.", "clpid": "Chiu-Daniel-T" }, { "family_name": "Huck", "given_name": "Wilhelm T. S.", "clpid": "Huck-W-T-S" }, { "family_name": "Kenis", "given_name": "Paul J. A.", "clpid": "Kenis-P-J-A" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Whitesides", "given_name": "George M.", "orcid": "0000-0001-9451-2442", "clpid": "Whitesides-G-M" } ], "abstract": "This Letter reports the measurement of electro-osmotic flows (EOF) in microchannels with surface charge patterned on the 200 mu m scale. We have investigated two classes of patterns: (1) Those in which the surface charge varies along a direction perpendicular to the electric field used to drive the EOF; this type of pattern generates multidirectional flow along the direction of the field. (2) Those in which the surface charge pattern varies parallel to the field; this pattern generates recirculating cellular flew, and thus causes motion both parallel and perpendicular to the external field. Measurements of both of these flours agree well with theory in the Limit of thin double layers and low surface potential.", "doi": "10.1103/PhysRevLett.84.3314", "issn": "0031-9007", "publisher": "American Physical Society", "publication": "Physical Review Letters", "publication_date": "2000-04-10", "series_number": "15", "volume": "84", "issue": "15", "pages": "3314-3317" }, { "id": "authors:z555n-g6v03", "collection": "authors", "collection_id": "z555n-g6v03", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160717870", "type": "article", "title": "Fabrication of metallic microstructures using exposed, developed silver halide-based photographic film", "author": [ { "family_name": "Deng", "given_name": "Tao", "clpid": "Deng-Tao" }, { "family_name": "Arias", "given_name": "Francisco", "clpid": "Arias-F" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Kenis", "given_name": "Paul J. A.", "clpid": "Kenis-P-J-A" }, { "family_name": "Whitesides", "given_name": "George M.", "orcid": "0000-0001-9451-2442", "clpid": "Whitesides-G-M" } ], "abstract": "This paper demonstrates that the pattern of silver particles embedded in the gelatin matrix of exposed and developed silver halide-based photographic film can serve as a template in a broadly applicable method for the microfabrication of metallic microstructures. In this method, a CAD file is reproduced in the photographic film by exposure and developing. The resulting pattern of discontinuous silver grains is augmented and made electrically continuous by electroless deposition of silver, and the electrically continuous structure is then used as the cathode for electrochemical deposition of an additional layer of the same or different metal. The overall process can be completed within 2 h, starting from a CAD file, and can generate electrically continuous structures with the smallest dimension in the plane of the film of 30 \u03bcm. Structures with aspect ratio of up to 5 can also be obtained by using the metallic structures as photomasks in photolithography using SU-8 photoresist on the top of the electroplated pattern and exposed from the bottom, followed by development and electroplating through the patterned photoresist. This method of fabrication uses readily available equipment and makes it possible to develop prototypes of a wide variety of metallic structures and devices. The resulting structures either supported on the film backing or freed from it are appropriate for use as passive, structural materials such as wire frames or meshes and can also be used in microfluidic, microanalytical, and microelectromechanical systems.", "doi": "10.1021/ac991010p", "issn": "0003-2700", "publisher": "American Chemical Society", "publication": "Analytical Chemistry", "publication_date": "2000-02-15", "series_number": "4", "volume": "72", "issue": "4", "pages": "645-651" }, { "id": "authors:0c10a-0z069", "collection": "authors", "collection_id": "0c10a-0z069", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160720579", "type": "article", "title": "Experimental and theoretical scaling laws for transverse diffusive broadening in two-phase laminar flows in microchannels", "author": [ { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Stroock", "given_name": "Abraham D.", "clpid": "Stroock-A-D" }, { "family_name": "Kenis", "given_name": "Paul J. A.", "clpid": "Kenis-P-J-A" }, { "family_name": "Whitesides", "given_name": "George", "orcid": "0000-0001-9451-2442", "clpid": "Whitesides-G-M" }, { "family_name": "Stone", "given_name": "Howard A.", "clpid": "Stone-H-A" } ], "abstract": "This letter quantifies both experimentally and theoretically the diffusion of low-molecular-weight species across the interface between two aqueous solutions in pressure-driven laminar flow in microchannels at high Peclet numbers. Confocal fluorescent microscopy was used to visualize a fluorescent product formed by reaction between chemical species carried separately by the two solutions. At steady state, the width of the reaction-diffusion zone at the interface adjacent to the wall of the channel and transverse to the direction of flow scales as the one-third power of both the axial distance down the channel (from the point where the two streams join) and the average velocity of the flow, instead of the more familiar one- half power scaling which was measured in the middle of the channel. A quantitative description of reaction-diffusion processes near the walls of the channel, such as described in this letter, is required for the rational use of laminar flows for performing spatially resolved surface chemistry and biology inside microchannels and for understanding three-dimensional features of mass transport in shearing flows near surfaces.", "doi": "10.1063/1.126351", "issn": "0003-6951", "publisher": "American Institute of Physics", "publication": "Applied Physics Letters", "publication_date": "2000", "series_number": "17", "volume": "76", "issue": "17", "pages": "2376-2378" }, { "id": "authors:bakgz-9e982", "collection": "authors", "collection_id": "bakgz-9e982", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160726623", "type": "article", "title": "Temperature effects on electron transfer within intervalence bis(hydrazine) radical cations", "author": [ { "family_name": "Nelsen", "given_name": "Stephen F.", "clpid": "Nelsen-S-F" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Gentile", "given_name": "Kevin E.", "clpid": "Gentile-K-E" }, { "family_name": "Powell", "given_name": "Douglas R.", "clpid": "Powell-D-R" } ], "abstract": "Analyses of the shape of intervalence charge-transfer bands at various temperatures (255\u2212325 K in most cases) in acetonitrile are reported for the radical cations of bis(2-tert-butyl-2,3-diazabicyclo[2.2.2]oct-3-yl) hydrazines that are bridged by 2,5-xylene-1,4-diyl (2^+), durene-1,4-diyl (3^+), naphthalene-1,4-diyl (4^+), biphenyl-4,4'-diyl (5^+), and 9,9-dimethylfluorene-2,7-diyl (6^+) aromatic rings. Electron-transfer (ET) rate constants measured by ESR as a function of temperature are reported for 4^+\u22126^+. Despite the fact that the ET barriers for these compounds are dominated by vibrational reorganization, explicit inclusion of vibronic coupling effects is not necessary for the prediction of their ET rate constants from the optical spectra. Rate constants in excellent agreement with the measured ones are predicted by a classical analysis of charge-transfer band shape, if the diabatic surfaces are changed from the usual assumption that they are parabolas to ones that fit the shape of the charge-transfer bands.", "doi": "10.1021/ja984047k", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "1999-08-04", "series_number": "30", "volume": "121", "issue": "30", "pages": "7108-7114" }, { "id": "authors:8ztzk-xej66", "collection": "authors", "collection_id": "8ztzk-xej66", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160726092", "type": "article", "title": "Ion pairing effects on bis(hydrazine) intervalence radical cations", "author": [ { "family_name": "Nelsen", "given_name": "Stephen F.", "clpid": "Nelsen-S-F" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Three bis(2-tert-butyl-2,3-diazabicyclo[2.2.2]oct-3-yl) radical cation salts, bridged by 2,3,5,6-tetramethylbenzene-1,4-diyl (1^+PF_6^-), biphenylene-4,4'-diyl (2^+PF_6^-), and 9,9-dimethyl-fluorene-2,7-diyl (3^+NO_3^-) groups, have been studied in methylene chloride. The transition energy at band maximum (E_(op)) increases as concentration increases and when ^nBu_4+BF_6^- is added, indicating that ion pairing increases E_(op). The E_(op) data fit a simple ion pairing equilibrium, giving ion pairing equilibrium constants at 293 K of 3100, 3100, and 6100 M^(-1), respectively. Electron-transfer rate constants measured by ESR are reported for 0.19 mM 2^+PF_6^- and for 1 mM 2^+PF_6^- and 3^+NO_3^- in the presence of 20 mM ^nBu_4+BF_6^- in methylene chloride. Prediction of k_(ET) from the optical spectrum of 2^+PF_6^- containing excess ^nBu_4^+BF_6^- was made both assuming the optical ET is endoenthalpic by an amount calculated from the increase in E_(op), and that \u0394G\u00b0 = 0 (that is, that the ion pairing effect may be lumped into the electron transfer coordinate along with the vertical and solvent reorganization effects). The predicted rate constant for the latter is only a factor of 2.5 times larger the former, so both agree rather well with the ESR-derived rate constant.", "doi": "10.1021/jp991294h", "issn": "1089-5639", "publisher": "American Chemical Society", "publication": "Journal of Physical Chemistry A", "publication_date": "1999-07-08", "series_number": "27", "volume": "103", "issue": "27", "pages": "5373-5378" }, { "id": "authors:xg3ce-ad362", "collection": "authors", "collection_id": "xg3ce-ad362", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160722004", "type": "article", "title": "Microfabrication inside capillaries using multiphase laminar flow patterning", "author": [ { "family_name": "Kenis", "given_name": "Paul J. A.", "clpid": "Kenis-P-J-A" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Whitesides", "given_name": "George M.", "orcid": "0000-0001-9451-2442", "clpid": "Whitesides-G-M" } ], "abstract": "The reaction of species in solutions flowing laminarly (without turbulent mixing) inside capillaries was used as the basis for a broadly applicable method of microfabrication. In this method, patterning occurs as a result of transport of reactive species to interfaces within the capillary by laminar flow. A wide range of chemistries can be used to generate structures with feature sizes of less than 5 micrometers and with spatial localization to within 5 micrometers. The method is applicable to the patterning of metals, organic polymers, inorganic crystals, and ceramics on the inner walls of preformed capillaries, using both additive and subtractive processes.", "doi": "10.1126/science.285.5424.83", "issn": "0036-8075", "publisher": "American Association for the Advancement of Science", "publication": "Science", "publication_date": "1999-07-02", "series_number": "5424", "volume": "285", "issue": "5424", "pages": "83-85" }, { "id": "authors:5bzbc-s3b95", "collection": "authors", "collection_id": "5bzbc-s3b95", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160733567", "type": "article", "title": "Patterning cells and their environments using multiple laminar fluid flows in capillary networks", "author": [ { "family_name": "Takayama", "given_name": "Shuichi", "clpid": "Takayama-Shuichi" }, { "family_name": "McDonald", "given_name": "J. Cooper", "clpid": "McDonald-J-Cooper" }, { "family_name": "Ostuni", "given_name": "Emanuele", "clpid": "Ostuni-E" }, { "family_name": "Liang", "given_name": "Michael N.", "clpid": "Liang-Michael-N" }, { "family_name": "Kenis", "given_name": "Paul J. A.", "clpid": "Kenis-P-J-A" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Whitesides", "given_name": "George M.", "orcid": "0000-0001-9451-2442", "clpid": "Whitesides-G-M" } ], "abstract": "This paper describes the use of laminar flow of liquids in capillary systems to pattern the cell culture substrate, to perform patterned cell deposition, and to pattern the cell culture media. We demonstrate the patterning of the cell culture substrate with different proteins, the patterning of different types of cells adjacent to each other, the patterned delivery of chemicals to adhered cells, and performing enzymatic reactions over select cells or over a portion of a cell. This method offers a way to simultaneously control the characteristics of the surface to which cells are attached, the type of cells that are in their vicinity, and the kind of media that cells or part of a cell are exposed to. The method is experimentally simple, highly adaptable, and requires no special equipment except for an elastomeric relief that can be readily prepared by rapid prototyping.", "doi": "10.1073/pnas.96.10.5545", "pmcid": "PMC21896", "issn": "0027-8424", "publisher": "National Academy of Sciences", "publication": "Proceedings of the National Academy of Sciences of the United States of America", "publication_date": "1999-05-11", "series_number": "10", "volume": "96", "issue": "10", "pages": "5545-5548" }, { "id": "authors:ekp2c-92x14", "collection": "authors", "collection_id": "ekp2c-92x14", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160735218", "type": "article", "title": "Complexity in chemistry", "author": [ { "family_name": "Whitesides", "given_name": "George M.", "orcid": "0000-0001-9451-2442", "clpid": "Whitesides-G-M" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "\"Complexity\" is a subject that is beginning to be important in chemistry. Historically, chemistry has emphasized the approximation of complex nonlinear processes by simpler Linear ones. Complexity is becoming a profitable approach to a wide range of problems, especially the understanding of Life.", "doi": "10.1126/science.284.5411.89", "issn": "0036-8075", "publisher": "American Association for the Advancement of Science", "publication": "Science", "publication_date": "1999-04-02", "series_number": "5411", "volume": "284", "issue": "5411", "pages": "89-92" }, { "id": "authors:4a5jf-4h871", "collection": "authors", "collection_id": "4a5jf-4h871", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160733739", "type": "article", "title": "Intra- and inter-molecular exchange on symmetrical hydrazine diradical dications and comparison of the magnetic exchange with ET parameters derived from their optical spectra", "author": [ { "family_name": "Teki", "given_name": "Yoshio", "clpid": "Teki-Y" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Nelsen", "given_name": "Stephen F.", "clpid": "Nelsen-S-F" } ], "abstract": "The spin alignment in a charged molecular field is important research issue in the molecular magnetism. In order to clarify the interrelation between spin alignment and the charged molecular field, we have investigated intra- and inter-molecular exchanges on some Hydrazine diradical dictations 1\u20133 shown in Scheme 1 (see texts) by ESR and magnetic susceptibility measurement. The magnetic behavior of the dication salt 1 has been well analyzed using the alternating linear chain models with J_(intra)/k_B = \u2212106 K, J_(inter_/k_B = \u221249 K and an alternating parameter \u03b1=0.46. The magnetic property of 3 has been also fitted to the alternating chain model with J/k_B = \u2212106 K J_(inter)/k_B = \u221242 K (\u03b1=0.40). On the other hand, 2 gives a robust triplet ground state with larger energy separation from other spin states. The energy separation has been estimated to be larger than 300 cm\u22121 (J_(intra)/k_B > \u00b1190 K) from the temperature dependence of the ESR signal intensity. These findings indicate that the sign of the intramolecular exchange depends on the linking position (m- or p-) of the hydrazine cation group, i.e. the topology of the \u03c0 orbital network even in the cationic molecular field. The magneto-optical correlation is also discussed based on the intramolecular electron transfer (ET) parameters.", "doi": "10.1080/10587259908023329", "issn": "1542-1406", "publisher": "Taylor & Francis", "publication": "Molecular Crystals and Liquid Crystals Science and Technology. Section A. Molecular Crystals and Liquid Crystals", "publication_date": "1999", "series_number": "1", "volume": "334", "issue": "1", "pages": "313-322" }, { "id": "authors:tp393-cqa46", "collection": "authors", "collection_id": "tp393-cqa46", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160726422", "type": "article", "title": "Indirect determination of self-exchange electron transfer rate constants", "author": [ { "family_name": "Nelsen", "given_name": "Stephen F.", "clpid": "Nelsen-S-F" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Gentile", "given_name": "Kevin E.", "clpid": "Gentile-K-E" }, { "family_name": "Nagy", "given_name": "Mark A.", "clpid": "Nagy-M-A" }, { "family_name": "Tran", "given_name": "Hieu Q.", "clpid": "Tran-Hieu-Q" }, { "family_name": "Qu", "given_name": "Qinling", "clpid": "Qu-Qinling" }, { "family_name": "Halfen", "given_name": "DeWayne T.", "clpid": "Halfen-D-T" }, { "family_name": "Odegard", "given_name": "Amy L.", "clpid": "Odegard-A-L" }, { "family_name": "Pladziewicz", "given_name": "Jack R.", "clpid": "Pladziewicz-J-R" } ], "abstract": "Second-order rate constants kij(obsd) measured at 25 \u00b0C in acetonitrile by stopped-flow spectrophotometry for forty-four electron transfer (ET) reactions among fourteen 0/+1 couples [three aromatic compounds (tetrathiafulvalene, tetramethyltetraselenafulvalene, and 9,10-dimethyl-9,10-dihydrophenazine), four 2,3-disubstituted 2,3-diazabicyclo[2.2.2]octane derivatives, six acyclic hydrazines, and the bridgehead diamine 1,5-diazabicyclo[3.3.3]undecane] and seventeen compounds and forty-seven reactions from a previous study (J. Am. Chem. Soc. 1997, 119, 5900) [three p-phenylenediamine derivatives, four ferrocene derivatives, and ten tetraalkylhydrazines] are discussed. When all 91 kij(obsd) values are simultaneously fitted to Marcus's adiabatic cross rate formula kij(calcd) = (kiikjjKijfij)1/2, ln fij = (ln Kij)2/4 ln(kiikjj/Z2), best-fit self-exchange rate constants, kii(fit), are obtained that allow remarkably accurate calculation of kij(obsd); kij(obsd)/kij(calcd) is in the range 0.5\u22122.0 for all 91 reactions. The average difference without regard to sign, |\u0394\u0394Gij|, between observed cross reaction activation free energy and that calculated using the kii(fit) values and equilibrium constants is 0.13 kcal/mol. The \u0394Gii(fit) values obtained range from 2.3 kcal/mol for tetramethyltetraselenafulvalene0/+ to 21.8 kcal/mol for tetra-n-propylhydrazine0/+, corresponding to a factor of 2 \u00d7 1014 in kii(fit). The principal factor affecting kii(fit) for our data appears to be the internal vertical reorganization energy (\u03bbv), but kii(fit) values also incorportate the effects of changes in the electronic matrix coupling element (V). Significantly smaller V values for ferrocenes and for hydrazines with alkyl groups larger than methyl than for aromatics and tetramethylhydrazine are implied by the observed \u0394Gii(fit) values.", "doi": "10.1021/ja9810890", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "1998-08-19", "series_number": "32", "volume": "120", "issue": "32", "pages": "8230-8240" }, { "id": "authors:ap9t0-9h954", "collection": "authors", "collection_id": "ap9t0-9h954", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160727245", "type": "article", "title": "Effects of bridge redox state levels on the electron transfer and optical properties of intervalence compounds with hydrazine charge-bearing units", "author": [ { "family_name": "Nelsen", "given_name": "Stephen F.", "clpid": "Nelsen-S-F" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Powell", "given_name": "Douglas R.", "clpid": "Powell-D-R" } ], "abstract": "n/a", "doi": "10.1021/ja960500l", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "1998-07-03", "series_number": "8", "volume": "120", "issue": "8", "pages": "1924-1925" }, { "id": "authors:mgkx8-3bn18", "collection": "authors", "collection_id": "mgkx8-3bn18", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160719423", "type": "article", "title": "Mathematical description of kinetic resolution with an enantiomerically impure catalyst and nonracemic substrate", "author": [ { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" } ], "abstract": "Kinetic resolution is an important method in organic chemistry; catalytic kinetic resolution is especially attractive because a smaller amount of the optically active material is required. In principle, a naturally occurring catalyst (enzyme) or a synthetic catalyst can be employed. Possible disadvantages of an enzymatic catalyst include limited scope of reactions and substrates; therefore, there is an increasing effort targeted at the design of chemical catalysts for kinetic resolution. The most useful parameter in comparing different catalysts is the selectivity factor S, which is the ratio of the rate constants for the reaction of the catalyst with the two enantiomers of the substrate (S = (k_1/k_2), see below). Mathematical treatment of kinetic resolution should provide a way to calculate S from experimental observables, and for enantiomerically pure catalysts the equations are well-known from the work of Kagan and others.", "doi": "10.1021/jo972299i", "issn": "0022-3263", "publisher": "American Chemical Society", "publication": "Journal of Organic Chemistry", "publication_date": "1998-05-29", "series_number": "11", "volume": "63", "issue": "11", "pages": "3772-3774" }, { "id": "authors:w22j8-dze88", "collection": "authors", "collection_id": "w22j8-dze88", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160728363", "type": "article", "title": "Structural information from hydrazine radical cation optical absorption spectra", "author": [ { "family_name": "Nelsen", "given_name": "Stephen F.", "clpid": "Nelsen-S-F" }, { "family_name": "Tran", "given_name": "Hieu Q.", "clpid": "Tran-Hieu-Q" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Ramm", "given_name": "Michael T.", "clpid": "Ramm-M-T" }, { "family_name": "Chen", "given_name": "Ling-Jen", "clpid": "Chen-Ling-Jen" }, { "family_name": "Powell", "given_name": "Douglas R.", "clpid": "Powell-D-R" } ], "abstract": "Transition energies (Eop) of the nitrogen-centered \u03c0,\u03c0* absorption of tetraalkylhydrazine radical cations are quite sensitive to twist at the NN bond, nitrogen pyramidality, and mixing of the \u03c3 orbitals with the \u03c0 system. Thirty-one examples for which Eop varies from 63 to 107.5 kcal/mol are discussed with the aid of calculated values (Ecalc) for the 0,0 transition energy using simple (no configuration interaction) neutral-in-cation-geometry calculations on AM1\u2212UHF geometry-optimized radical-cation structures. Significant changes in the difference between Eop and Ecalc are observed for bis-N,N'-bicyclic systems, which are syn pyramidalized at nitrogen (twist angles near 0\u00b0; Eop about 23 kcal/mol larger than Ecalc) and for bis-N,N-bicyclic ones, which are anti pyramidalized (twist angles of 180\u00b0; difference about 7 kcal/mol when calculations of 180\u00b0 structures are employed). Within these classes, changes in Eop caused by changes in pyramidality and \u03c3,\u03c0 interaction are predicted well by the calculations. The tetraisopropylhydrazine radical cation has \u03bbmax = 282 nm, but its tetracyclohexyl analogue shows two transitions, at 276 and 386 nm. This surprising difference is attributed to tetracyclohexylhydrazine radical cation having both untwisted and significantly twisted (estimated twist angle \u2248 44\u00b0) forms occupied in solution, although the isopropyl compound only has the untwisted form significantly occupied.", "doi": "10.1021/jo9718307", "issn": "0022-3263", "publisher": "American Chemical Society", "publication": "Journal of Organic Chemistry", "publication_date": "1998-04-17", "series_number": "8", "volume": "63", "issue": "8", "pages": "2536-2543" }, { "id": "authors:p0nt0-a3k69", "collection": "authors", "collection_id": "p0nt0-a3k69", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160730344", "type": "article", "title": "Conformational analysis of a 12-membered crown dithioether in the solid state and in solution", "author": [ { "family_name": "Samoshin", "given_name": "Vyacheslav V.", "clpid": "Samoshin-V-V" }, { "family_name": "Troyansky", "given_name": "Emmanuil I.", "clpid": "Troyansky-E-I" }, { "family_name": "Demchuk", "given_name": "Dmitry V.", "clpid": "Demchuk-D-V" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Chertkov", "given_name": "Vyacheslav A.", "clpid": "Chertkov-V-A" }, { "family_name": "Lindeman", "given_name": "Sergei V.", "clpid": "Lindeman-S-V" }, { "family_name": "Khrustalyov", "given_name": "Victor N.", "clpid": "Khrustalyov-V-N" }, { "family_name": "Struchkov", "given_name": "Yury T.", "clpid": "Struchkov-Y-T" } ], "abstract": "The solid-state molecular structure and the conformational behaviour in solution of the 12-membered crown dithioether 8- methyl-1,4-dioxa-7,10-dithiacyclododecane-5,12-dione were studied by x-ray crystallography, H-1 and C-13 NMR spectroscopy and molecular mechanics. The conformational rigidity of some constituent structural fragments allowed a detailed analysis of the structure and distribution of the conformers. A protocol for studies of multiconformational equilibrium was developed by means of the combined use of structure calculations and dynamic NMR measurements. (C) 1998 John Wiley & Sons, Ltd.", "doi": "10.1002/(SICI)1099-1395(199804)11:4<241::AID-POC999>3.0.CO;2-X", "issn": "0894-3230", "publisher": "Wiley", "publication": "Journal of Physical Organic Chemistry", "publication_date": "1998-04", "series_number": "4", "volume": "11", "issue": "4", "pages": "241-253" }, { "id": "authors:k3953-8sv75", "collection": "authors", "collection_id": "k3953-8sv75", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160727396", "type": "article", "title": "Comparison of the singlet, triplet energy gap of a symmetrical diradical dication with ET parameters derived from its optical spectrum", "author": [ { "family_name": "Nelsen", "given_name": "Stephen F.", "clpid": "Nelsen-S-F" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Teki", "given_name": "Yoshio", "clpid": "Teki-Y" } ], "abstract": "n/a", "doi": "10.1021/ja9706615", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "1998-03-11", "series_number": "9", "volume": "120", "issue": "9", "pages": "2200-2201" }, { "id": "authors:6jtfg-dvc59", "collection": "authors", "collection_id": "6jtfg-dvc59", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160727624", "type": "article", "title": "Adiabatic electron transfer: Comparison of modified theory with experiment", "author": [ { "family_name": "Nelsen", "given_name": "Stephen F.", "clpid": "Nelsen-S-F" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Trieber", "given_name": "Dwight A., II", "clpid": "Trieber-D-A-II" } ], "abstract": "The radical cations of properly designed bishydrazines allow comparison of observed and calculated electron transfer rate constants. These compounds have rate constants small enough to be measured by dynamic electron spin resonance spectroscopy and show charge transfer bands corresponding to vertical excitation from the energy well for the charge occurring upon one hydrazine unit to that for the electron-transferred species. Analysis of the data for all six compounds studied indicates that the shape of the adiabatic surface on which electron transfer occurs can be obtained from the charge transfer band accurately enough to successfully predict the electron transfer rate constant and that explicit tunneling corrections are not required for these compounds.", "doi": "10.1126/science.278.5339.846", "issn": "0036-8075", "publisher": "American Association for the Advancement of Science", "publication": "Science", "publication_date": "1997-10-31", "series_number": "5339", "volume": "278", "issue": "5339", "pages": "846-849" }, { "id": "authors:y9edv-x4885", "collection": "authors", "collection_id": "y9edv-x4885", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160727091", "type": "article", "title": "Charge-localized p-phenylenedihydrazine radical cations: ESR and optical studies of intramolecular electron transfer rates", "author": [ { "family_name": "Nelsen", "given_name": "Stephen F.", "clpid": "Nelsen-S-F" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Powell", "given_name": "Douglas R.", "clpid": "Powell-D-R" } ], "abstract": "1,4-Bis(2-tert-butyl-2,3-diazabicyclo[2.2.2]oct-3-yl)benzene-1,4-diyl (2) its 2,5-dimethyl and 2,3,5,6-tetramethyl derivatives (3 and 4), their radical cations, and bis-radical dications are studied. Crystal structures including those of 2^+BPh_4^-, 3^(2+)(BPh_4^-)_2, 4^+BPh_4^-, and 4^(2+)(BPh_4^-)_2 establish that ring methylation causes more N-lone pair, aryl \u03c0 twist without changing the NAr,NAr distance significantly and that both 2^+ and 4^+ have the charge localized in one hydrazine unit. NMR measurements show that 3^+ has about 6% of its spin at the four aryl CH and CMe carbons, while 4^+ has about 1.5% of its spin at the four CMe carbons. The average distance between the unpaired electrons of 3^(2+) and 4^(2+) was obtained from the dipolar splittings of their thermally excited triplet states and, as expected, is significantly smaller for 3^(2+) (5.25 \u00c5) than for 4^(2+) (5.63 \u00c5). Rate constants for electron transfer between the hydrazine units of 3^+ and 4^+ in CH_2Cl_2 and CH_3CN were determined by dynamic ESR. The intervalence radical cations show charge transfer bands corresponding to vertical electron transfer between the ground state and the highly vibrationally excited electron-shifted state, allowing calculation of the parameters controlling electron transfer. Electron transfer parameters obtained from the CT bands using adiabatic energy surfaces which approximate the CT band shapes observed produce rate constants within experimental error of those extrapolated to room temperature from the ESR data for both 3^+ and 4^+ in both solvents, without using tunneling corrections. The effects of mixing of the electronic wave functions of the reduced and oxidized hydrazine units of 2^+ on d_(NN), the C(t-Bu)N,NA(Ar) twist angle, and the aryl nitrogen lone pair, aryl \u03c0 twist angle which are observed by X-ray are close to those predicted from the position of the minima on the ET coordinate X of the adiabatic energy surface calculated from the CT band.", "doi": "10.1021/ja9720321", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "1997-10-22", "series_number": "42", "volume": "119", "issue": "42", "pages": "10213-10222" }, { "id": "authors:sv9a6-acy48", "collection": "authors", "collection_id": "sv9a6-acy48", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160727814", "type": "article", "title": "\u03c3,\u03c0 Interaction in Halogen-Substituted Biadamantylidene Radical Cations", "author": [ { "family_name": "Nelsen", "given_name": "Stephen F.", "clpid": "Nelsen-S-F" }, { "family_name": "Klein", "given_name": "Susan J.", "clpid": "Klein-S-J" }, { "family_name": "Trieber", "given_name": "Dwight A., II", "clpid": "Trieber-D-A-II" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Powell", "given_name": "Douglas R.", "clpid": "Powell-D-R" } ], "abstract": "The order of E\u00b0' and vIP for 4-eq-halogenated-biadamantylidene is F > Cl Br, and the 5-F-substituted compound is harder to ozidize than the 4-eq-F-substituted one. The former result is most consistent with a detectable resonance contribution through the \u03c3-framework, and the latter with \u03c3-hyperconjugative destablilization proceeding through two pathways being more than double the same effect through one pathway (the Whiffen effect). AM1 calculations predict these results. The facial selectivity for epoxidation and diazetidine formation from 4-eq-halogenated 3 (4(X)) is in the order Cl > F > Br, and the 5-fluoro compound (8) is less selective than 4(F) for both reactions. Steric as well as electronic factors might well contribute to these results, neither of which was expected from consideration of \u03c3,\u03c0 interaction. Cation radical catalyzed chain dioxetane formation from 4(F) and 3(Cl) is significantly more face selective than epoxidation or diazetidine formation, as expected on electronic grounds; \u03c3,\u03c0 interaction should be larger in the radical cation.", "doi": "10.1021/jo970252r", "issn": "0022-3263", "publisher": "American Chemical Society", "publication": "Journal of Organic Chemistry", "publication_date": "1997-09-19", "series_number": "19", "volume": "62", "issue": "19", "pages": "6539-6546" }, { "id": "authors:ae9h4-dtp61", "collection": "authors", "collection_id": "ae9h4-dtp61", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160728161", "type": "article", "title": "Estimation of self-exchange electron transfer rate constants for organic compounds from stopped-flow studies", "author": [ { "family_name": "Nelsen", "given_name": "Stephen F.", "clpid": "Nelsen-S-F" }, { "family_name": "Ramm", "given_name": "Michael T.", "clpid": "Ramm-M-T" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Nagy", "given_name": "Mark A.", "clpid": "Nagy-M-A" }, { "family_name": "Trieber", "given_name": "Dwight A., II", "clpid": "Trieber-D-A-II" }, { "family_name": "Powell", "given_name": "Douglas R.", "clpid": "Powell-D-R" }, { "family_name": "Chen", "given_name": "Xi", "clpid": "Chen-Xi-Wisconsin-Eau-Claire" }, { "family_name": "Gengler", "given_name": "Jamie J.", "clpid": "Gengler-J-J" }, { "family_name": "Qu", "given_name": "Qinling", "clpid": "Qu-Qinling" }, { "family_name": "Brandt", "given_name": "Jennifer L.", "clpid": "Brandt-J-L" }, { "family_name": "Pladziewicz", "given_name": "Jack R.", "clpid": "Pladziewicz-J-R" } ], "abstract": "Second-order rate constants k12(obsd) measured at 25 \u00b0C in acetonitrile by stopped-flow for 47 electron transfer (ET) reactions among ten tetraalkylhydrazines, four ferrocene derivatives, and three p-phenylenediamine derivatives are discussed. Marcus's adiabatic cross rate formula k12(calcd) = (k11 k22 k12 f12)1/2, ln f12 = (ln K12)2/4 ln(k11k22/Z2) works well to correlate these data. When all k12(obsd) values are simultaneously fitted to this relationship, best-fit self-exchange rate constants, kii(fit), are obtained that allow remarkably accurate calculation of k12(obsd); k12(obsd)/k12'(calcd) is in the range of 0.55\u22121.94 for all 47 reactions. The average \u0394\u0394Gij between observed activation free energy and that calculated using kii(fit) is 0.13 kcal/mol. Simulations using Jortner vibronic coupling theory to calculate k12 using parameters which produce the wide range of kii values observed predict that Marcus's formula should be followed even when V is as low as 0.1 kcal/mol, in the weakly nonadiabatic region. Tetracyclohexylhydrazine has a higher kii than tetraisopropylhydrazine by a factor of ca. 10. Replacing the dimethylamino groups of tetramethyl-p-phenylenediamine by 9-azabicyclo[3.3.1]nonyl groups has little effect on kii, demonstrating that conformations which have high intermolecular aromatic ring overlap are not necessary for large ET rate constants. Replacing a \u03b3 CH2 group of a 9-azabicyclo[3.3.1]nonyl group by a carbonyl group lowers kii by a factor of 17 for the doubly substituted hydrazine and by considerably less for the doubly substituted p-phenylenediamine.", "doi": "10.1021/ja970321j", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "1997-06-25", "series_number": "25", "volume": "119", "issue": "25", "pages": "5900-5907" }, { "id": "authors:pva1b-de575", "collection": "authors", "collection_id": "pva1b-de575", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160734554", "type": "article", "title": "Homolytic cycloaddition of dithiols to alkynes leading to sulfur-containing heterocycles and crown thioethers: new comments to the mechanism", "author": [ { "family_name": "Troyansky", "given_name": "Emmanuil I.", "clpid": "Troyansky-E-I" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Samoshin", "given_name": "Vyacheslav V.", "clpid": "Samoshin-V-V" } ], "abstract": "Multicenter three-electron (3e-2c) S-S bond formation can be employed to explain the cyclization in the homolytic addn. of \u03b1,\u03c9-thiols to alkynes.", "issn": "0278-6117", "publisher": "Harwood", "publication": "Sulfur Letters", "publication_date": "1997", "series_number": "5", "volume": "20", "issue": "5", "pages": "219-224" }, { "id": "authors:rem7e-rcw77", "collection": "authors", "collection_id": "rem7e-rcw77", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160726929", "type": "article", "title": "Charge localization in a dihydrazine analogue of tetramethyl-p-phenylenediamine radical cation", "author": [ { "family_name": "Nelsen", "given_name": "Stephen F.", "clpid": "Nelsen-S-F" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Powell", "given_name": "Douglas R.", "clpid": "Powell-D-R" } ], "abstract": "n/a", "doi": "10.1021/ja960500l", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "1996-07-03", "series_number": "26", "volume": "118", "issue": "26", "pages": "6313-6314" }, { "id": "authors:tx32h-adc58", "collection": "authors", "collection_id": "tx32h-adc58", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160726256", "type": "article", "title": "Slow electron transfer reactions involving tetraisopropylhydrazine", "author": [ { "family_name": "Nelsen", "given_name": "Stephen F.", "clpid": "Nelsen-S-F" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Chen", "given_name": "Ling-Jen", "clpid": "Chen-Ling-Jen" }, { "family_name": "Brandt", "given_name": "Jennifer L.", "clpid": "Brandt-J-L" }, { "family_name": "Chen", "given_name": "Xi", "clpid": "Chen-Xi-Wisconsin-Eau-Claire" }, { "family_name": "Pladziewicz", "given_name": "Jack R.", "clpid": "Pladziewicz-J-R" } ], "abstract": "n/a", "doi": "10.1021/ja9537080", "issn": "0002-7863", "publisher": "American Chemical Society", "publication": "Journal of the American Chemical Society", "publication_date": "1996-02-14", "series_number": "6", "volume": "118", "issue": "6", "pages": "1555-1556" }, { "id": "authors:3nc7s-fm496", "collection": "authors", "collection_id": "3nc7s-fm496", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160734712", "type": "article", "title": "Stereoselective Free-Radical Cycloaddition-Macrocyclization in Facile Synthesis of Trans-Cyclohexano-Fused 12-Membered Crown Thioethers", "author": [ { "family_name": "Troyansky", "given_name": "Emmanuil I.", "clpid": "Troyansky-E-I" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Samoshin", "given_name": "Vyacheslav V.", "clpid": "Samoshin-V-V" }, { "family_name": "Strelenko", "given_name": "Yury A.", "clpid": "Strelenko-Y-A" }, { "family_name": "Demchuk", "given_name": "Dmitry V.", "clpid": "Demchuk-D-V" }, { "family_name": "Nikishin", "given_name": "Gennady I.", "clpid": "Nikishin-G-I" }, { "family_name": "Lindeman", "given_name": "Sergey V.", "clpid": "Lindeman-S-V" }, { "family_name": "Khrustalyov", "given_name": "Viktor N.", "clpid": "Khrustalyov-V-N" }, { "family_name": "Struchkov", "given_name": "Yury T.", "clpid": "Struchkov-Y-T" } ], "abstract": "Homolytic cycloaddition of dithiols 1,2 derived from trans- and eis-1,2-cyclohexanediols to alkynes, induced by Pr3B-O2, offers an extremely simple approach to trans- and cis-cyclohexano-fused 12-membered crown thialactones 4a-c-7a-c. The reaction of trans-1 proceeds with pronounced remote 1,6-asymmetric induction to give predominantly (IS *, 6R *, 12S *)-4a\u2013c, while cis-2 reacts nonstereoselectively. Basing on molecular mechanics calculations the stereoselectivity is rationalized as a result of entropy favored pathway of macrocyclization.", "doi": "10.1016/0040-4020(95)00707-F", "issn": "0040-4020", "publisher": "Elsevier", "publication": "Tetrahedron", "publication_date": "1995-10-16", "series_number": "42", "volume": "51", "issue": "42", "pages": "11431-11444" }, { "id": "authors:t6b6k-s7795", "collection": "authors", "collection_id": "t6b6k-s7795", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160734870", "type": "article", "title": "Remote Asymmetric Induction in Free-Radical Cycloaddition Leading to Trans-Cyclohexano-Fused 12-Membered Crown Thioethers", "author": [ { "family_name": "Troyansky", "given_name": "Emmanuil I.", "clpid": "Troyansky-E-I" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Strelenko", "given_name": "Yury A.", "clpid": "Strelenko-Y-A" }, { "family_name": "Samoshin", "given_name": "Vyacheslav V.", "clpid": "Samoshin-V-V" }, { "family_name": "Demchuk", "given_name": "Dmitry V.", "clpid": "Demchuk-D-V" }, { "family_name": "Nikishin", "given_name": "Gennady I.", "clpid": "Nikishin-G-I" }, { "family_name": "Lindeman", "given_name": "Sergey V.", "clpid": "Lindeman-S-V" }, { "family_name": "Khrustalyov", "given_name": "Viktor N.", "clpid": "Khrustalyov-V-N" }, { "family_name": "Struchkov", "given_name": "Yury T.", "clpid": "Struchkov-Y-T" } ], "abstract": "Homolytic cycloaddition of dithiol 1, derived from trans-1,2-cyclohexanediol, to alkynes induced by Pr3B\ue5f8O2 occurs with 1,6-asymmetric induction to afford predominantly (1S\u2217, 6R\u2217, 12S\u2217)-trans-cyclohexano-fused 12-membered crown thialactones 4a-c. No pronounced diasteroselectivity was found in the corresponding reactions of dithiol 2, derived from cis-1,2-cyclohexanediol.", "doi": "10.1016/0040-4039(95)00287-5", "issn": "0040-4039", "publisher": "Elsevier", "publication": "Tetrahedron Letters", "publication_date": "1995-03-27", "series_number": "13", "volume": "36", "issue": "13", "pages": "2293-2294" }, { "id": "authors:0qkn1-h1484", "collection": "authors", "collection_id": "0qkn1-h1484", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160717675", "type": "article", "title": "Synthesis of 12-Membered and 13-Membered Sulfur-Containing Lactones by Homolytic Macrocyclization of Mercaptoacetic Esters with Alkynes", "author": [ { "family_name": "Demchuk", "given_name": "Dmitry V.", "clpid": "Demchuk-D-V" }, { "family_name": "Lazareva", "given_name": "Margarita I.", "clpid": "Lazareva-M-I" }, { "family_name": "Lindeman", "given_name": "Sergey V.", "clpid": "Lindeman-S-V" }, { "family_name": "Khrustalyov", "given_name": "Viktor N.", "clpid": "Khrustalyov-V-N" }, { "family_name": "Struchkov", "given_name": "Yurii T.", "clpid": "Struchkov-Y-T" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Troyansky", "given_name": "Emmanuil I.", "clpid": "Troyansky-E-I" }, { "family_name": "Nikishin", "given_name": "Gennady I.", "clpid": "Nikishin-G-I" } ], "abstract": "Starting from mercaptoacetic acid esters of 1,2- or 1,3-diols and substituted acetylenes 12- and 13-membered sulfur-containing lactones as 1:1 adducts were synthesized in yields up to 48%. The mechanism of this homolytic reaction, which is initiated by the tripropylborane/oxygen system, includes generation of thiyl radicals and their addition to the triple bond of alkynes.", "doi": "10.1055/s-1995-3891", "issn": "0039-7881", "publisher": "Georg Thieme Verlag", "publication": "Synthesis-Stuttgart", "publication_date": "1995-03", "series_number": "3", "volume": "1995", "issue": "3", "pages": "307-311" }, { "id": "authors:ypz31-m0t46", "collection": "authors", "collection_id": "ypz31-m0t46", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160734243", "type": "article", "title": "An Approach to 8-Membered, 16-Membered and 24-Membered Sulfur- Containing Heterocycles Via Homolytic Cycloaddition of Alkynes with Butane-1,4-Dithiol", "author": [ { "family_name": "Troyansky", "given_name": "Emmanuil I.", "clpid": "Troyansky-E-I" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Korneeva", "given_name": "Ekaterina N.", "clpid": "Korneeva-E-N" }, { "family_name": "Pogosyan", "given_name": "Mariam S.", "clpid": "Pogosyan-M-S" }, { "family_name": "Nikishin", "given_name": "Gennady I.", "clpid": "Nikishin-G-I" } ], "abstract": "Homolytic cycloaddition of butane-1,4-dithiol with alkynes offers a facile one-pot route to 8-membered 1,4-dithiocanes and 16-and 24-membered crown thioethers, 1,4,9,12-tetrathiacyclohexadecanes and 1,4,9,12,17,20-hexathiacyclotetracosanes.", "doi": "10.1070/MC1995v005n01ABEH000441", "issn": "0959-9436", "publisher": "Elsevier", "publication": "Mendeleev Communications", "publication_date": "1995", "series_number": "1", "volume": "5", "issue": "1", "pages": "18-20" }, { "id": "authors:xwgef-ghj91", "collection": "authors", "collection_id": "xwgef-ghj91", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20130821-160734409", "type": "article", "title": "Homolytic Macrocyclization of Alkynes with Propane-1,3-Dithiol as a Route to 14-Membered and 21-Membered Crown Thioethers", "author": [ { "family_name": "Troyansky", "given_name": "Emmanuil I.", "clpid": "Troyansky-E-I" }, { "family_name": "Demchuk", "given_name": "Dmitry V.", "clpid": "Demchuk-D-V" }, { "family_name": "Ismagilov", "given_name": "Rustem F.", "orcid": "0000-0002-3680-4399", "clpid": "Ismagilov-R-F" }, { "family_name": "Lazareva", "given_name": "Margarita I.", "clpid": "Lazareva-M-I" }, { "family_name": "Strelenko", "given_name": "Yurii A.", "clpid": "Strelenko-Y-A" }, { "family_name": "Nikishin", "given_name": "Gennady I.", "clpid": "Nikishin-G-I" } ], "abstract": "Crown thioethers, 1,4,8,11-tetrathiacyclotetradecanes and 1,4,8,11,15,18-hexathiaheneicosanes, have been synthesized as 2:2 and 3:3 cycloadducts in a 'one-pot' homolytic macrocyclization of alkynes with propane-1,3-dithiol.", "doi": "10.1070/MC1993v003n03ABEH000248", "issn": "0959-9436", "publisher": "Elsevier", "publication": "Mendeleev Communications", "publication_date": "1993", "series_number": "3", "volume": "3", "issue": "3", "pages": "112-114" } ]