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Isotachophoretic fluorescence in situ hybridization of intact bacterial cells
journal contribution
posted on 2017-06-20, 00:00 authored by Sui C Phung, Joan M Cabot, Mirek Macka, Shane M Powell, Rosanne GuijtRosanne Guijt, Michael BreadmoreA counter-pressure-assisted capillary isotachophoresis method in combination with a sieving matrix and ionic spacer was used to perform in-line fluorescence in situ hybridization (FISH) of bacterial cells. A high concentration of sieving matrix (1.8% w/v HEC) was introduced at one end of the capillary, and the bacterial cells were suspended in the spacer electrolyte for injection. Using a 2 min injection with 18 psi counter-pressure, 50% of the cells injected into the capillary were hybridized with the fluorescently labeled oligonucleotide, and the excess unhybridized probe was separated from the hybridized cell-probe complexes in a two-stage ITP method. With an LOD (6.0 × 104 cells/mL) comparable with the CE analysis of a sample processed using an off-line FISH protocol, the total analysis time was reduced from 2.5 h to 30 min. Provided the appropriate probe is selected, this approach can be used for specific detection of bacterial cells in aqueous samples.
History
Journal
Analytical chemistryVolume
89Issue
12Pagination
6513 - 6520Publisher
American Chemical SocietyLocation
Washington, D.C.Publisher DOI
ISSN
0003-2700eISSN
1520-6882Language
engPublication classification
C Journal article; C1.1 Refereed article in a scholarly journalCopyright notice
2017, American Chemical SocietyUsage metrics
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No categories selectedKeywords
fluorescence in situ hybridization (FISH)bacterial cellscapillaryoligonucleotideScience & TechnologyPhysical SciencesChemistry, AnalyticalChemistryFIELD-AMPLIFIED CONDITIONSCAPILLARY-ELECTROPHORESISELECTROKINETIC INJECTIONDIMETHYL-SULFOXIDEDNA HYBRIDIZATIONIDENTIFICATIONPROBESDIAGNOSTICSCULTUREASSAY
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