I need to urgently analyze some samples this week, but I have not yet antibodies. They take 2 or 3 weeks to arrive, so I want to know if I can keep the fixed cells. Will there be any change?
I would rather freeze the cells (-80 degree) in serum (about 50%) and DMSO (5-10%) containing medium. You can easily thaw and stain the cells weeks later.
If you do not want to freeze them, you can fix the cells with methanol (-20 degree) and store them for several weeks before staining, Be careful during staining and perform proper washing, because methanol will also permeabilize the cells.
Thanks for your reply. Does the process of thawing have any particular process or the procedure is similar to common thawing of cells?
Yes, usual thawing process should be okay. It will help to have some DNAse (10 ng/ml) in the FACS buffer along with EDTA (1-2 mM) and FBS (1-2%). DNA released from dead cells can result in cell clumps.
I suggest you make a validation study before you rely on freeze/thaw procedures. CD4 is more stable in comparison to CD8. You may use 5 - 10% DMSO in cell culture medium for freezing. After thawing, if you keep the cells at a refrigerator overnight you will have better results.
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