Phosphatase-Protease inhibitors in TCA total protein extraction - (Nov/03/2008 )
I am using TCA/acetone protocol for total protein extraction. I was wondering if there is any use for protease/phosphatase inhibitors, when the first step is grounding in liquid nitrogen and then precipitation with acetone. Peopole in my lab use both PMSF and phosphatase inhibitors coctail, but isn't it usless?
Thanks for your suggestions
When i have resolubilized TCA precipitate, I add protease inhibitors. If your protein is phosphorylated or kinase sensitive, then i would use phosphatase inhibitors. I have found that Roche cocktail complete inhibitors work well.
Thanks, so I was probably right. I don't think I can add inhibitors to solubilization - it is last step before IEF and inhibitors would interfer with focusation, but I will think about it.