I use 4 % PFA to fix my cells on a coverslip. How long should the cells be treated with PFA? Also, how should I maintain these fixed slides for longer periods? Are these slides stored with some liquid to avoid drying?
Hi,
10 minutes at room temperature works well, should not incubate more than 15 minutes. Usually, fixation is done just before staining. Therefore, it always better after staining and sealing you could preserve at 4-degree Celcius for imaging purposes.
10 min for cells is enough, if you want more time, leave in refrigerator.
10 minutes works well but depending on cell type you may need to go a little longer. As for storage, I would keep them in PBS in the refrigerator. However, remember that in a buffer, even fixed cells undergo extraction over time. So as for storage I would limit it to 2 or 3 days at most, depending upon what you are planning to do with them.
Fixing in paraformaldehyde for more than 10-15 minutes will cross link the proteins to the point where antigen retrieval may be required to ensure the antibody has free access to bind and detect the protein.
Around 15-20 mins incubation in PFA works well for fixing cells. After fixation, you can store these slides at +4C in PBS for few days.
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