Source: Flaherty, R. A., et al. Implementation of a Permeable Membrane Insert-based Infection System to Study the Effects of Secreted Bacterial Toxins on Mammalian Host Cells. J. Vis. Exp. (2016).
This video demonstrates a permeable membrane insert system to investigate the impact of bacterial toxins on host cells in a controlled and isolated manner. This approach allows for the selective exposure of the cells to the toxins while maintaining a physical separation that mimics the natural host-pathogen interaction.
1. Bacterial Culture
2. Host Cell Culture
3. Host Cell Infection with Permeable Membrane Insert System
4. Sample Collection
The authors have nothing to disclose.
HaCaT human epithelial keratinocytes | Gift from lab of Victor Nizet. | ||
Dulbecco's Modified Eagle's Medium (DMEM) | Life Technologies | 11995-073 | Use appropriate media for cell line of interest. |
Fetal Bovine Serum (FBS) | Biowest | S162H | Use appropriate media additives for cell line of interest. |
10 cm tissue culture dishes | Nunc | 150350 | Other brands are suitable. |
6-well tissue culture dishes | CytoOne | cc7682-7506 | Other brands are suitable but need to be compatible with the Corning insert. |
Collagen-coated 0.4 μm Transwell inserts (6 well) | Corning | 3540 | |
Collagen-coated 0.4 μm Transwell inserts (24 well) | Corning | 3595 | |
Cell scrapers | Fisherbrand | 08-100-241 | These are only required for the collection of cell lysates. |
Nonidet P40 Substitute | Sigma | 74385-1L | Other suppliers are suitable. |