Re: Cromogen/Substrate IHC question

From:Sandra Livingston <slivings@hscprime.hsc.usf.edu>


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Victoria, I am wondering if you meant to suggest that Catherine try the
Vector NovaRed that works with the ABC-HRP system instead of switching
to the Vector Red which works with the ABC-Alk-Phos.

Catherine, unless there is some specific reason for switching to Vector
Red (prefer that shade of red? want to do double staining with a
different systems?), I agree with Victoria about simply switching to a
red chromogen with the HRP system.  Unlike AEC, the NovaRed CAN be taken
through the alcohols and xylene and mounted with a resinous mounting
medium without appreciable loss of signal if the slides are not allowed
to sit for any length of time in the alcohols.  I also agree that it is
best to either re-titrate your reagents (if you feel you must change to
Alk Phos) or just run a couple of antibodies that you know work well on
known positives with both the ABC-HRP and ABC-Alk Phos at the standard
dilutions and compare the staining.

Victoria Baker wrote:

> Hi,
>
> When switching detection systems it is always safer to check your
> dilutions again and do a titration run.  But instead of switching
> this, why not get the red substrate that Vector has for Alk-Phos?  The
> kit is like $65.00 and will save more in time an effort on your part.
> I keep both systems in my lab (HRP & Alk Phos) with red chromogen.
> We've had good results for the most part.
>
> Good luck.
>
> Vikki Baker
> American Health Foundation
> Valhalla, New York 10595
>
>
>   "Bennett, Catherine (Katie)" <cbennett@lrri.org> wrote:
>
>      Advice needed:
>
>      If I switch an IHC procedure over from an ABC-HRP kit to an
>      ABC-Alk Phos
>      kit, I shouldn't have to adjust my primary and secondary
>      antibody
>      concentrations, right? (I want to change cromogen labels
>      from DAB to
>      Vector-Red.)
>
>      TIA.
>
>      *********************************
>      Catherine "Katie" Bresee Bennett
>      Sr. Research Technologist
>      Lovelace Respiratory Research Institute
>      Albuquerque, New Mexico
>
>
>
> -----------------------------------------------------------------------
> Do You Yahoo!?
> Yahoo! Auctions - Buy the things you want at great prices.

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<!doctype html public "-//w3c//dtd html 4.0 transitional//en">
<html>
Victoria, I am wondering if you meant to suggest that Catherine try the
Vector NovaRed that works with the ABC-HRP system instead of switching
to the Vector Red which works with the ABC-Alk-Phos.
<p>Catherine, unless there is some specific reason for switching to Vector
Red (prefer that shade of red? want to do double staining with a different
systems?), I agree with Victoria about simply switching to a red chromogen
with the HRP system.  Unlike AEC, the NovaRed CAN be taken through
the alcohols and xylene and mounted with a resinous mounting medium without
appreciable loss of signal if the slides are not allowed to sit for any
length of time in the alcohols.  I also agree that it is best to either
re-titrate your reagents (if you feel you must change to Alk Phos) or just
run a couple of antibodies that you know work well on known positives with
both the ABC-HRP and ABC-Alk Phos at the standard dilutions and compare
the staining.
<p>Victoria Baker wrote:
<blockquote TYPE=CITE>Hi,
<p>When switching detection systems it is always safer to check your dilutions
again and do a titration run.  But instead of switching this, why
not get the red substrate that Vector has for Alk-Phos?  The kit is
like $65.00 and will save more in time an effort on your part.  I
keep both systems in my lab (HRP & Alk Phos) with red chromogen. 
We've had good results for the most part.
<p>Good luck.
<p>Vikki Baker
<address>
American Health Foundation</address>

<address>
Valhalla, New York 10595</address>

<p><br>  <b><i>"Bennett, Catherine (Katie)" <cbennett@lrri.org></i></b>
wrote:
<blockquote style="BORDER-LEFT: #1010ff 2px solid; MARGIN-LEFT: 5px; PADDING-LEFT: 5px">Advice
needed:
<p>If I switch an IHC procedure over from an ABC-HRP kit to an ABC-Alk
Phos
<br>kit, I shouldn't have to adjust my primary and secondary antibody
<br>concentrations, right? (I want to change cromogen labels from DAB to
<br>Vector-Red.)
<p>TIA.
<p>*********************************
<br>Catherine "Katie" Bresee Bennett
<br>Sr. Research Technologist
<br>Lovelace Respiratory Research Institute
<br>Albuquerque, New Mexico
<br> </blockquote>

<p><br>
<hr size=1><b>Do You Yahoo!?</b>
<br><a href="http://auctions.yahoo.com">Yahoo! Auctions</a> - Buy the things
you want at great prices.</blockquote>
</html>

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