I want to determine if my peptide induces survival on transfected CHO K1 cells, and i want to use H2O2 as apoptotic agent. I verify apoptosis by double staining propidium iodide/annexin V with flow cytometry.
hi,
see this paper more details about H2O2 concentration induce apoptosis
Hi,
Depends on incubation time and quality of your cells.
I would suggest to make a quick dose-effect test in any case.
In vitro, 50 to 100 uM are considered moderate doses with moderate to mild effects. 200 uM to 400 uM are considered high and sublethal doses (but it also depends on incubation duration).
500 uM is a typical dose with which you begin to see obvious cell death; high H2O2 oxidize cysteine thiols and destroy enzyme activity.
0.5 mM of H2O2 is an ideal concentration that would induce cell death for 3 hours. H2O2 induces both necrosis and apoptosis.
I made a dose response for 1h incubation and 24h, but it is very variable, CHO-K1 can be very resistant. And i want to very if my cells can to survive with a peptide pretreatment so i don't want treat my cells with H2O2 for a long time.
I try with 3 hours incubation.
YOU MUST CHECK THE MORPHOLOGY OF CELLS EXPOSED FOR 24 h with . h2 02 before peptide treatment
May apoptosis dose and duration of hydrogen peroxide vary from epithelial cell line to squamus cell line ?
I also want to induce apoptosis in 293T cell line through H2O2 and check the expression of particular protein expression after and before treatment of the molecule which can inhibit or downregulate the apoptosis of the cells. Is it possible to see this process through apoptosis induced by H2O2?
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