[Histonet] Can DAB block an epitope??

[C.M. van der Loos]

Herbert and Rene,To my opinion DAB is the only know chromogen reaction product that does have the potential to block an epitope. Afterall, the paper of Sternberger and Joseph (JHC 1979) describes a double staining procedure with two primaries of similar species based on DAB as first chromogen, no elution step or heating step, and subsequent performance of a 2nd IHC procedure. That protocol works perfect as long as DAB is used in the first staining sequence. This double staining procedure is however NOT working with other chromogens. If other chromgens are used, a heating step is required to remove the immuno-reagents from the first staining sequence. My paper in JOH of last year (31:119-127) combines for example two AP procedures (red and blue) for double staining. Cheers,ChrisChris van der Loos, PhD
Dept. of Pathology
Academic Medical Center M2-230
Meibergdreef 9
NL-1105 AZ Amsterdam
The Netherlands Date: Fri, 14 Aug 2009 09:55:58 -0700 (PDT)
From: Rene J Buesa <>
Subject: Re: [Histonet] Can DAB block an epitope??
To: Histonet <@t> lists.utsouthwestern.edu, GT Hebert
emerald_lake77 <@t> yahoo.com
Theoretically, and depending on the reaction specificity, DAB should not block the epitope.
Ren� J.

--- On Fri, 8/14/09, GT Hebert <emerald_lake77 <@t> yahoo.com> wrote:


From: GT Hebert <emerald_lake77 <@t> yahoo.com>
Subject: [Histonet] Can DAB block an epitope??
To: Histonet <@t> lists.utsouthwestern.edu
Date: Friday, August 14, 2009, 11:37 AM


Hello all,
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I was hoping to run a chromogenic immuno to check if two proteins colocalize.� 
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Since both antibodies are made in rabbit, I was going to stain one and visualize with DAB and then heat retrieve (pressure or steam) to drop off the first antibody and restain with my second rabbit antibody using a different color to visualize.
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How likely, if at all will the DAB precipitate block / mask the epitope of my second antibody if indeed they are in a similar location?
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Any info would be greatly appreciated.
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Thank you!
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Regards.
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Gustave
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Gustave T. Hebert
Research Scientist I
Metabolic Disease Research
Wyeth Research
200 CambridgePark Drive
Cambridge, MA 02140