Re: [Histonet] staining mouse tissues using mouse monoclonals

From:

Jackie.O'Connor@abbott.com

I routinely use mouse primaries on Mouse tissue using either the ARK kit (K3954) from DAKO.
I've tried the M.O.M. from Vector (they have a couple of 'flavors') and the MM Biotinylation Kit from Biocare (MMBK-G).
All are equally superb for mouse on mouse, so take your pick. DAKO has a handy little Excel spreadsheet you can download for making the dilutions for the ARK -it comes in really handy, and you can print out the dilution sheet for data retention.

Jacqueline M. O'Connor HT(ASCP)
Abbott Laboratories
Global Pharmaceutical Research and Development
Discovery Chemotheraputics

ALAN MEEKER <ameeker@mail.jhmi.edu>
Sent by: histonet-admin@lists.utsouthwestern.edu

10/28/2003 12:25 PM

To: histonet@lists.utsouthwestern.edu
cc:
Subject: [Histonet] staining mouse tissues using mouse monoclonals

Hi, I am trying immunohistochem on mouse tissues using a mouse IgG monoclonal. The target is nuclear, and I can see the signals fairly well. The problem is in background staining from the endogenous mouse antibody, especially in things like plasma cells in the lamina propria of the large intestine. I am wondering what is the best way to attenuate or eliminate this background signal.

I tried something simple, namely pre-treating the tissue with an excess of goat anti-mouse IgG, hoping to bind/cover up most of the resident IgG before applying my primary and then HRP-anti mouse secondary. This seems not to have worked at all, perhaps because there were unused binding sites on the goat anti-mouse that bound my monoclonal primary (?).

At any rate, the simple, quick fix didn't do the trick. I have heard that there are "mouse on mouse" kits available to deal with this problem. Has anyone had any experience using these?

-Alan Meeker

Alan Meeker, PhD
Department of Pathology
Division of Genitourinary Pathology
Bunting-Blaustein Cancer Research Building Room 153
1650 Orleans Street
Baltimore, MD 21231-1000
e-mail: ameeker@mail.jhmi.edu

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