Isotachophoretic Fluorescence in Situ Hybridization of Intact Bacterial Cells

Analytical Chemistry
Sui C PhungMichael C Breadmore

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.

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Citations

Oct 12, 2017·Lab on a Chip·C Eid, J G Santiago
Jun 12, 2018·Biomicrofluidics·D Huber, G V Kaigala
Jul 25, 2018·Electrophoresis·Zdena Malá, Petr Gebauer
Nov 21, 2019·Nanoscale·Dan LiVincent Rotello
Jan 4, 2018·Analytical Chemistry·Robert L C VoetenGovert W Somsen
Nov 28, 2021·Electrophoresis·Monica N AlvesMichael C Breadmore

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