I try to culture PBMC from whole blood Without any growth stimulation. I found a problem that within one hour the culture appeared like contaminated. I checked the same with different individuals blood samples. I couldn’t realise what is the actual contamination.
Dear Sheethal,
I don't think that you can detect any contamination in your cultures only 1 hour after isolation; I think that you are seeing platelets. To double-check, leave the cells in the incubator for couple of days and check if they are really contaminated.
I hope it helps!
Hello Sheethal S Kumar
After the PBMC layer is harvested, which has to be done by careful pipetting, you need to wash the cells twice with PBS or cell medium (RPMI-1640) in order to remove the platelets and the residual separation medium.
Is the washing step included in the protocol?
Best.
Dear Malcolm thank you for your response. we wash the cells twice with PBS and then suspend in RPMI media. we observe only lymphocytes in a clear background. Just after 1 hour we observed some dark particles larger than the cells.
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