The caspase-8 may cleave RIP3, how to detect the tRIP3?? Is there any antibody or other assay to measure it??
I have not seen a RIP3 antibody that specifically detect the cleaved form. The best is to use the good antibodies that detect full length RIP3 and have controls with and without z-VAD.fmk or Boc-D.fmk (general caspase inhibitors). You should see a reduction in RIP3 levels in the absence of the caspase inhibitor (if caspase-8 is indeed cleaving RIP3 in your model), while in the presence of the caspase inhibitor, the RIP3 band should be similar to that of untreated control.
Good luck with your studies.
It would depend on the system. In a cell culture system Dr. Samali's suggestion would be an indirect way to see whether RIP3 is being cleaved. To detect the cleavage product: Labelled recombinant protein (labelled at C-terminus) might be a way to do it in vitro. Few papers suggest that RIP3 ab's are poor at identifying the endogenous cleaved fragment - however there are several antibodies and some might work.
Don't have a copy with me. Please do a search on pubmed and several manuscripts should be free and probably your institution has a subscription for a lot of them too.
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