PI staining - (Oct/17/2006 )
the data is so confusing. I would go back to the begining.
First I treated my cells with cytokine or control, MTT data indicated cytokine inhibits cell proliferation.
Second, I use PI staining to confirm this. but results showed 1) no apoptosis (no sub-g1 peak)，2) less cells in G1 by using cytokine, 3) same cells in s, and 4) more cells in G2/M by using cytokine. so PI staining result is contradictory to MTT assay.
Third, as recommanded by you, I detected the expression of CycB1, it is up regulated by cytokine.
So, do you have some idea? which data I can believe or how to explain it?
BTW, when I perform MTT assay, all readings are within linear range, so I think MTT results reflected cell number accurately and results are very reproducible.
Thanks and look forward to your help.
When you use PI staining method for cell cycle analysis (including subG1, G1, S/G2/M), do you also collect the medium and pellet the possible cells in the medium for analysis? My purpose is to detect how many cells in apoptosis, will apoptotic cell detach from the dish?
What is your PI Satining method/protocol?
I am also interested in studying cell cycle differences in my cells under different types of stresses like mitochondrial stress, oxidative stress (H2O2), etc