[Histonet] re: Background on FFPE mouse tissue
gcallis <@t> montana.edu
Mon Sep 17 12:54:26 CDT 2007
Good point, rat antimouse, hamster antimouse, or rabbit antimouse (if they
have a monoclonal for mouse made in rabbit) or rabbit antihuman.
Not only secondaries adsorbed to mouse, but also use mouse serum in
diluents/normal serum blocking. We prefer F(ab')2 frag of IgG secondaries,
adsorbed to mouse in order to eliminate fc receptor binding. Jackson has
excellent antibodies for that purpose.
Many of the kits for clinical use contain multilinks for mouse, rabbit
and goat. If you use a single link polymer kit for rodents, goat, rabbit,
you will eliminate biotin and the mouse on mouse problems. Biocare Promark
single links work very well since they have an array of single links useful
to research. I believer Vision Biosystems also has this available.
If it is a mouse on mouse problem and you must use a mouse monoclonal, then
kits that biotinylate the primary are superb. DAKO ARK and Biocare have
excellent kits for this purpose. A lady gave an excellent poster on this
at NSH a few years back, and her results were very convincing for the use
of this kind of kit, no secondary antibodies are used at all.
At 11:17 AM 9/17/2007, you wrote:
>As Paul suggested, my first thought is that the background is due to the
>use of mouse monoclonals. A mouse-on-mouse kit may help, but my limited
>experience with these has not been very positive; they may be a bit
>unpredictable, and/or cause a marked decrease in the signal. Much
>preferable is to use primary antibodies from another species.
>Jean-Martin Lapointe, DMV, MS, dACVP
>Histonet mailing list
>Histonet <@t> lists.utsouthwestern.edu
Research Histopathology Supervisor
Veterinary Molecular Biology
Montana State University - Bozeman
PO Box 173610
Bozeman MT 59717-3610
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