Dear Irit, Seems to me your AB does not bind to protein G as desired, and AB +target binds better.
You might try covalent cross-linking instead of protein G. There are e.g. carboxyl activated (magnetic) beads available. The coupling is easy: buffer exchange your AB by gel filtration into e.g. borate buffer, incubate with activated beads for 2hrs, quench and block, then wash with your target buffer. Another option is biotinylating your AB and using streptavidin coated beads. Procedure is similar as above. Concerning BSA as control, I'd prefer an unrelated antibody of the same IgG subclass instead. Should have less possible "side effects". HTH Wo _______________________________________________ Methods mailing list [email protected] http://www.bio.net/biomail/listinfo/methods
