stripping western blot - (Oct/09/2006 )
Im using westerns at the minute to detect two proteins (20, 100kda). Do i have to strip the blot and reprobe or is it possible to use both primaries at the same time?
If you are sure that your two antibodies give only one band, you can incubate both in the same time. If not, detect first one protein, then the second one. You don't need to strip.
you can also cut the blot between the two proteins and incubate separately the two half-blots.
missele is right; moreover, it should be tested if each antibody does not crossreact with polypeptides in the ranges of interest, f.i. the Ab against 20 kDa should not crossreact with a polypeptide in the rane of 100 kDa, and vice versa for the other Ab; if there are crossreactions you should not use them in parallel
The only potenital problem maybe the resolution. You may need to run different percentage gels in order to get a clean product.