immunofluorescence background

Yasuhito Abe yasuhito at ME1SERV.M.EHIME-U.AC.JP
Sat Mar 18 02:19:45 EST 1995


At  3:54 AM 95.3.17 -0800, Dave Gilbert wrote:
>Does anyone have any tricks to reduce apparently non-specific association
>of polyclonal antibodies.  We have tried blocking steps with serum,
>however, our immunofluorescent signals with antibody are still only
>marginally above signals obtained with pre-immune serum.
>
>Thanks.
>
>Dave Gilbert
>Department of Biochemistry and Molecular Biology
>SUNY Health Science Center
>750 E. Adams St.
>Syracuse, N.Y. 13210
>Tel:(315) 464-8723
>FAX:(315) 464-8750


Try adsorption of your antibody (usually IgG) using pre-immue serum: ex.
using rabbit serum attached to Sepharose 4B.  If it did not work well, try
to prepare F(ab)2 fragment instead of whole IgG.  If it were still not
good, you should use affinity purified antibody using antigen-attached
column such as Sepharose 4B-something.  The first thing is high population
and high avidity of specific antibody in your immune-serum, and the next
thing is fighting against rheumatic factor in its broad meaning.  Good
luck.
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#       Yasuhito Abe, M.D.                       #
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