Detection of Bacterial Adherence to Host Cells Using Fluorescence Imaging

Published: October 31, 2023

Abstract

Source: Yang, J., et al. Automated, High-Throughput Detection of Bacterial Adherence to Host Cells. J. Vis. Exp. (2021)

This video demonstrates the bacterial adherence assay to evaluate GFP-tagged bacterial adherence to epithelial cell culture to study the dynamics of host-pathogen interactions using fluorescence microscopy-based imaging.

Protocol

1. Bacterial adherence and host cell staining First, wash the seeded A549 cell monolayers three times with warm 1x PBS. For each wash, add 100 μL of 1x PBS to each well, gently pipette up and down three times, dispose of 1x PBS or wait for 10 s after addition, and then vacuum to remove 1x PBS. To determine the kinetics of bacterial association, overlay the cells with 100 µL of desired concentrations of bacterial suspension with different MOIs (0, 1, 10, and 100). Spin down the bacte…

Divulgations

The authors have nothing to disclose.

Materials

10x PBS VWR 45001-130
4′,6-diamidino-2-phenylindole (DAPI) Thermo Fisher 62248 Host cell staining dye
96-well plate Corning 3882 Half area well, flat clear bottom
A549 cells ATCC CCL 185 Mammalian cell line
Centrifuge Eppendorf 5810R
OD600 DiluPhotometer IMPLEN
Typical soy broth Growcells MBPE-4040
F-12k medium ATCC  302004 A549 cell culture medium
Fetal bovine serum Corning 35-016-CV

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Detection of Bacterial Adherence to Host Cells Using Fluorescence Imaging. J. Vis. Exp. (Pending Publication), e21663, doi: (2023).

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