cross-linking antibodyies to beads
(by novalidaddress from nurfuerspam.de)
Tue Jan 19 02:02:15 EST 2010
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".
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