[Histonet] RE: Histonet Digest, Vol 81, Issue 1

[Perry, Margaret]

What antibody are you trying to use?

-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of histonet-request <@t> lists.utsouthwestern.edu
Sent: Sunday, August 01, 2010 12:01 PM
To: histonet <@t> lists.utsouthwestern.edu
Subject: Histonet Digest, Vol 81, Issue 1

Send Histonet mailing list submissions to
	histonet <@t> lists.utsouthwestern.edu

To subscribe or unsubscribe via the World Wide Web, visit
	http://lists.utsouthwestern.edu/mailman/listinfo/histonet
or, via email, send a message with subject or body 'help' to
	histonet-request <@t> lists.utsouthwestern.edu

You can reach the person managing the list at
	histonet-owner <@t> lists.utsouthwestern.edu

When replying, please edit your Subject line so it is more specific than "Re: Contents of Histonet digest..."


Today's Topics:

   1. Help with Biocare polymer detection kit!! (Jennifer Campbell)


----------------------------------------------------------------------

Message: 1
Date: Sat, 31 Jul 2010 12:03:39 -0700
From: "Jennifer Campbell" <jcampbell <@t> vdxpathology.com>
Subject: [Histonet] Help with Biocare polymer detection kit!!
To: <histonet <@t> lists.utsouthwestern.edu>
Message-ID:
	<5658CBDB9EAE6545ABE50D2563D81BF8181FF1 <@t> VDXSERVER01.vdxpathology.local>
	
Content-Type: text/plain;	charset="iso-8859-1"

I have been trying to get the Mach 1 polymer kit from Biocare to work on my K9 and feline tissue and I am getting terrible nonspecific staining in my positive and negative controls.  It appears to be sticking to something in the glands of the intestinal epithelia.  I am also getting nonspecific staining in my LN tissue.
I decided to switch over to polymer detection kit because I had been dealing with the issue of endogenous biotin when using the streptavidin biotin-based detection system, and now I am getting this other nonspecific staining!
I have tried doing runs eliminating my protein block and eliminating the negative control and primary antibody step altogether (to see if maybe the protein block or the diluent used in my negative control and antibody dilutions are causing the problem).  I have also reduced the incubation time of both the probe and polymer from 15 min and 30 min, respectively, to 10 min each.  I have tried cutting down my antigen retrieval time substantially as well.
 
I'm not sure what else could be causing this.  Any suggestions??
 
Thanks in advance,
 
Jennifer Campbell


------------------------------

_______________________________________________
Histonet mailing list
Histonet <@t> lists.utsouthwestern.edu
http://lists.utsouthwestern.edu/mailman/listinfo/histonet

End of Histonet Digest, Vol 81, Issue 1
***************************************