I'm trying to stain colon cancer cells with PI to see both G0/G1 peak and G/M peak. I can see the G0/G1 peak, but it's not as sharp as I think it should be and I can't even see the G2/M peak. My method is the following:
Collect cells with trypsin, aspirate media from cell pellet
Treat cells with staining solution that contains 5microgram/mL PI, 5 microgram/mL boiled RNAse A in PBS, sodium citrate (i forget concentration), and 0.1% Triton X-100.
I incubate cells with staining solution in dark at 37C for 10min
Then I add NaCl to solution (i forget how much, don't have protocol with me)
Any suggestions would be helpful, I'm really just want this to work.