I would like to assess the levels of p-AKT,p-p70s6k and BAD in cardiac myocytes by flow cytometric analysis. I would add my agonist +- inhibitor for 10 minutes and prepare the cells for analysis. Do you still need to look at total AKT, p70S6 and BAD levels too?
Daniel J. Medina
Hi Afthab,
I would include the total protein. Just like western blot, when ever you're looking at the phosphorylated form a protein it's important to include the total (non-phosphorylated) form. Besides, you never know what a reviewer is thinking. So it's much easier to include a few extra samples now than it is to wait for a review to come back and ask why you didn't look at total protein control.
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Mario Picozza
....Although 10 minutes is a short time for protein neosyntesis or degradation...
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