Hello All,
I am planing to do experiment on treating H2O2 (hydrogen peroxide) on SY5Y cell for 24 hr to determine concentration of different proteins, but before that I have to find the toxic dose of H2O2 from cell viability by MTT. I would like to know what is suitable toxic dose for determine protein signaling change in the cell. Many papers seems to select the dose that does not give lower than 50% cell viability. I am not sure whether there is a rule for that.
Thank you
The link Shared by Abdalla is excellent resource. I have refered it in past.
However, the actual concentration to be used must be tested by you in lab. I have seen signaling changes at 100 micromolar concentration as time as low as 10 min and up to 4 hr. However, time needs to be decided based on protein to be studied. For most antioxidant proteins 10-20 minutes is enough to see protein oxidation. However, for studying the change in expression, or transcription, or changes in downstream proteins in redox signaling, you should try 4-6 Hr treatment.
So far 4hrs looks best to me.
5mM H2O2 is sufficient to cause oxidative stress and its induced cell membrane damage in red blood cells.
Hi Asa,
I would use the ic10 from the MTT assay for cell signalling experiments, whereas the ic50 maybe better to observe failure of adaptation processes.
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