Immunoprecipitation in Non-reducing conditions? - Protocol suggestions? (Oct/04/2006 )
Is there was a way to do an immunoprecipitation under non-reducing conditions? My protocols have always involved boiling the beads in SDS-PAGE sample buffer with DTT, but was hoping we could liberate our antigen in some way without using the reducing agent. I would greatly appreciate any information you could supply!
-lil-
you may be able to release the antigen by changing the pH to 2.5 or 9.5 to 10 as you would for immunoaffinity chromatography.
-mdfenko-
Boiling the sample without DTT will separate the anitgen from the antibody, and the antibody from the beads.
The binding is non-covalent so SDS alone should destroy any bond formation.
-jaknight-