We want to do cd68 staining on human cell culture grow on coverslip. However, we are still waiting for the ordered antibody to arrive. So my question is, what is the best way to fix these cells and then store them so they can be stain later, like 1-2 month later?
Some methods i heard of are: 4%PFA & store in PBS, PFA & dry & store in -20degree, methanol & -20degree store, but don’t know which one is better for CD68.
thank you.
Dear Pam,
Sharing a link discussing about fixing adherent cells. People I know are fixing with methanol and it works well.
https://bitesizebio.com/13460/how-to-fix-adherent-cells-for-microscopy-and-imaging/
If you wanna fix cells for flowcytometry, then the cells have to be detached and stored in cryo medium.
You may want to consider the location of the antigen (extracellular or intracellular) since this is a transmembrane protein. Methanol will also permeabilize the cells and can interfere with downstream applications. I like to use 4% PFA and store in PBS at 4C (parafilm around the edges to avoid evaporation). Then, I can permeabilize them if needed during my staining protocol. I would not dry the coverslips.
For some cell lines, I have found that pre-fixing in 50/50 mix of 4% PFA/culture medium (so 2% PFA) improves preservation of cell morphology (and improved maintenance of cell attachment during staining).
You could try to fix them in Bouin´s fluid and than store them in 70% ethanol
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