Western Blotting with TransaBind

Anthony Shearing SHEARINA at QUCDN.QueensU.ca
Thu Jun 3 11:12:30 EST 1993


In article <MAILQUEUE-101.930603094516.16 at micro.uct.ac.za> Ed Rybicki,
ED at micro.uct.ac.za writes:
>Why bother?  Standard NC works fine, even for multiple elutions of
>antibody: I have done up to five elutions with only gradually diminishing
>yields, from proteins either Western blotted conventionally onto NC with
>no fixing, or simply from bits of NC soaked in the antigen, then
>washed and blocked.  A HELL of a lot cheaper, and FAR less fuss!!

I usually use NC for antibody purifications and it works well.  However,
the protein I am using is  hydrophobic and this is causing problems.  I
tried to do a blocking experiment depleting the antibody with membrane
bound protein but I  think a small amount of protein was released into
the antibody solution.  These proteins are then non-specifically binding
to western blotting membranes despite previous blocking.  The result is a
"negative" effect where the membrane itself is lighting up by ECL better
than bound protein.  I had hoped to use Transabind to covalently
immobilize the protein and prevent it from contaminating my antibody
solution.
Anthony Shearing
Shearina at QUCDN.queensu.ca
Queen's University
Ph. (613) 545-6424
Fax (613) 545-6830



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