westerns using monoclonals

nussbaum at neurochem.u-strasbg.fr nussbaum at neurochem.u-strasbg.fr
Wed Nov 2 03:48:43 EST 1994


In article <9410312005.AA14428 at phinet.smithkline.com> pattonaj%phvax.dnet at sb.com writes:
>
>I have a question along similar lines to that of QiWang.
>Is there anything you can do other than starting again if a monoclonal fails
>to immunoprecipitate or western blot? I used to work on a monoclonal that 
>gave a beautiful response on cryostat sections yet we could never do anything
>else with it. We eventually gave up but if there was anything I could try....
>
>Many thanks
>
>Amanda Patton


I have the same problem too working with a Mab directed against
a surface antigen of oligodendrocytes.
I suppose that your epitope is conformation dependent; so I suggest 
the following: extract your material with 1 or 2% CHAPS containing
 0.5 M NaCl and try a dot blotting research .You will already see if
 there is some immunoreaction; If it is positive my assumption is 
probably correct. After that you may perhaps try an affinity column
with this extract and it will probably work!!!
Let me know your results



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