[Histonet] Re: false positive kidney

From:"Alicia Henn Ph.D."

Hi Jackie,
    We struggled with similar background problems staining human kidney and 
liver. Blocking didn't help. We switched to the Dako Envision polymer-based 
system for detection, which is biotin-free. Problems solved.

Obi-wan. Now that's a name I've not heard in a long time...
Alicia Henn, Ph.D.
Vaccinex, Inc.

Date: Wed, 9 Mar 2005 16:32:48 -0600
From: "Jackie M O'Connor" 
Subject: [Histonet] It's a mystery . . .
To: Histonet@lists.utsouthwestern.edu

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Hi Histonetskis -
Maybe I'm thinking too hard - I can't seem to get rid of false positive
staining in negative control sections of human and mouse kidney human and
pancreas tumor.  I'm using EDTA HIER (for the first time), in an SABC
protocol with a Goat Pab.  I've tried two different (vendor) Rab anti Goat
biotinylated secondarys.   I'm using an avidin/biotin block - each for 20
minutes, (two different lots/vendors) but I'm still getting this false
staining in the negative (omitted primary) control sections.  Is it
possible to not be able to block ALL biotin? I've never seen this before.
I can't imagine what else I'm seeing - but like I said - my brain is
tired.  Before I switch to an HRP conjugated secondary - can anyone
suggest what else this could be?  I know EDTA can cause background, but
there is no primary on these sections.  Will someone loan me their brain?
I can't help but wonder if the EDTA is causing my mystery - but I have to
use it.
Help me, Obi-wan-kenobi.

Jackie O' 

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