how to get red of high secondary signal - (Jul/02/2011 )

hi,
i'm doing western and want to reprobe the membrane with different secondary ab the problem that I still get the previous signal also, I heard that you can block the signal using ponceau instead of stripping,do anyone know protocol?

ponceau is just a dye to stain proteins on the membrane, I have never heard it could actually strip it. I think you are better of redoing your western blot.

Are you using HRP or alkaline phosphatase as the secondary conjugate? If HRP, you should be able to use Sodium Azide to permanantly block the activity of the first secondary conjugate.

bob1 on Sat Jul 2 22:32:18 2011 said:

Basically just add some azide (i'd go for about 0.1% w/v) to some PBS and incubate for a couple of hours. In theory this should work, though I have never tested it myself. Azide is used to block endogenous peroxidase in IHC.