[Histonet] RE: Stupid rabbit antibodies

[Patsy Ruegg]

Are you using the rab abs on human samples or another species?  For most
purposes serum free protein block should be all you need.  If you were using
rab ab on rabbits I would recommend the FC block from Innogenex, you can
also get a anti rab link that is Fab2 and perhaps absorbed to the species
you are working with.  Southern Biotech sells really good secondary link
reagents, u can get it hrp, ap, fitc, or unlabeled and then match up the
detection to the animal the link is made in.  are you using a avidin/biotin
detection system?  If you are there may be endogenous biotin which can be
blocked.

Patsy

Patsy Ruegg, HT(ASCP)QIHC
IHCtech
12635 Montview Blvd. Ste.215
Aurora, CO 80045
720-859-4060
fax 720-859-4110
www.ihctech.net 
www.ihcrg.org


-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Perry,
Margaret
Sent: Thursday, March 24, 2011 1:05 PM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] RE: Stupid rabbit antibodies




Have you tried the Dako Rabbit envision?  We also tried the AbCam Expose
Rabbit specific HRP and used Nova red from Vector  or the DAB in the kit
depending on which antibody we used.  It is excellent , maybe even a little
better than Dako's envision.  I did have to use blocks such as FC receptor
block Innovex, Powerblock Biogenex and serum free protein block DAKO.  What
I used depended on the antibody with some needing only the block provided in
the kit.  Good Luck.

Margaret Perry HT(ASCP) 
Dept of Veterinary and  Biomedical services
Box 2175
South Dakota State University
Brookings SD 57007
605-688-5638


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Message: 17
Date: Tue, 22 Mar 2011 16:04:16 -0500
From: <sgoebel <@t> mirnarx.com>
Subject: [Histonet] Stupid Rabbit primaries!
To: <histonet <@t> lists.utsouthwestern.edu>
Message-ID:
	<D957F2A7D21959488C492A2680F9920A1C5FF5 <@t> svrexch.asuragen.us>
Content-Type: text/plain;	charset="us-ascii"

So I haven't had to deal with rabbit polyclonal primaries in a long time
because I remember how much the background sucks with them.
Unfortunately the only available antibody is a rabbit polyclonal.  Does
anyone have any suggestions for how to eliminate the background?  I have
diluted almost to the point of the antigens not showing!  Thanks guys
and gals!!

 

Sarah Goebel, BA, HT(ASCP)

Histotechnologist

Mirna Therapeutics

2150 Woodward Street

Suite 100

Austin, Texas  78744

(512)901-0900 ext. 6912

 



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

Message: 18
Date: Tue, 22 Mar 2011 16:13:04 -0500
From: "Sebree Linda A" <LSebree <@t> uwhealth.org>
Subject: RE: [Histonet] Stupid Rabbit primaries!
To: <sgoebel <@t> mirnarx.com>,	<histonet <@t> lists.utsouthwestern.edu>
Message-ID:
	
<8C023B4AB999614BA4791BAEB26E2738010029DE <@t> UWHC-MAIL01.uwhis.hosp.wisc.edu>
	
Content-Type: text/plain;	charset="us-ascii"

Do you know for sure that its the Ab and not endogenous biotin,
peroxidase or Phosphatase?  You can block for all those.

Otherwise there are a number of commercial blocking agents on the
market...try Biocare for a start (800)799-9499 or better yet, the
company that makes your antibody.

Good luck. 


Linda A. Sebree
University of Wisconsin Hospital & Clinics
IHC/ISH Laboratory
DB1-223 VAH
600 Highland Ave.
Madison, WI 53792
(608)265-6596



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