When I am performing Immunoflurescnce staining on fixed cells, should I work in the hood or it is ok to work on bench?
Thank you,
Dear Muhannad,
We use the hood, because after fixing in formaldehyde, or other fixing sol the cells are dead, but we try to keep our antibodies steriel, and dark for other in vivo studies.
Best,
It is only necessary to work in a cell culture hood to grow the cells before fixation. Once the cells have reached the end-point of an experiment, although you may perform fixation and IF staining in the hood, we normally bring the culture vessel (with cells growing on the sterile coverslip) out to an open bench to proceed with the remaining protocol steps. This way, you can spare the cell culture hood as there are various incubation times involved in IF staining and some steps would require ambient light sources or incubation in the dark, which are more accessible in areas outside of the cell culture hood.
You might also want to prepare the formaldehyde you fix your cells with in a fume hood as it is toxic, but the rest of the protocol may be carried out in an open bench, similar to immunohistochemistry (IHC) staining. We use a different brand of IF kit, but the protocol involved is fairly similar to a general protocol described in the file attached. Good luck!
Another Question, can I keep and reused the antibody that I added on cells before wash step?
Thank you.
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