I wish to perform PI staining on ishikawa cells for cell cycle analysis, would like to know the optimum amount of RNAse A to be added here for 1 million cells?
Usually 0.2-0.5 microgram RNase/ml is added to cells from a stock (usually 20 mg RNase A/ml). If cells are in ~1 ml solution, the range would be 10-25 microliter of the stock . After adding RNase A, I would incubate cells for about an hour at 37 degree C.
Working concentration of RNAse 40 microgram/microlitre for 1h at 37C incubation will be sufficient for PI staining.(Stock concentration of RNAase should be -1 mg/ml, and from there 5 microlitre is needed for 100 microlitre cell suspension volume).
Any advice for cell fixation and staining for flow cytometry? I was used to stain with DAPI for cell cicle analysis but naw I've a new flow cytometer that is not abble to dead DAPI.
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