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Isotachophoretic fluorescence in situ hybridization of intact bacterial cells

Phung, SC, Cabot, JM ORCID: 0000-0002-3305-078X, Macka, M ORCID: 0000-0002-6792-2574, Powell, SM ORCID: 0000-0001-5082-1630, Guijt, RM ORCID: 0000-0003-0011-5708 and Breadmore, M ORCID: 0000-0001-5591-4326 2017 , 'Isotachophoretic fluorescence in situ hybridization of intact bacterial cells' , Analytical Chemistry, vol. 89, no. 12 , pp. 6513-6520 , doi: 10.1021/acs.analchem.7b00598.

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Abstract

A 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.

Item Type: Article
Authors/Creators:Phung, SC and Cabot, JM and Macka, M and Powell, SM and Guijt, RM and Breadmore, M
Keywords: fluorescence in situ hybridization, bacterial cells,
Journal or Publication Title: Analytical Chemistry
Publisher: Amer Chemical Soc
ISSN: 0003-2700
DOI / ID Number: 10.1021/acs.analchem.7b00598
Copyright Information:

Copyright 2017 American Chemical Society

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