What should I add to the buffer after collagenase digestion? I'm now using 1% BSA/200 nM adenosine/PBS to wash the cells. After staining with DAPI and membrane marker I mix 1:10 with glycerol and embed onto slide. I would like to have the cells evenly spread on the slide. Any suggestions?
Does the PBS have calcium? It it a modified PBS or does it just have phosphate and saline? If it is a modified PBS, make a solution with reduced calcium. Otherwise, the addition of EDTA might be worth trying.
DNA strands released from cells (after a rather harsh dissociation) could clump the cells. In that case, DNAse would be helpful after digestion with collagenase.
I fully agree with Ratneswary, i have observed this in the thawed cord blood samples, where the cells have been frozen in high density Addition of DNAase helps in preventing the clumping, you can add it along with the enzyme during the digestion.
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