[Histonet] FW: Help!

[Patti Loykasek]

I  don't have experience with animal tissue, but can give my experiences
with human tissue. Every once in a while we will be asked to use a slide
that has already been heat pretreated & the slide is either a negative IHC
control or the original antibody was not positive. In that situation, we
remove the coverslip, rehydrate the slide, and DO NOT repeat the heat
retrieval. We have had positive staining with this method, but I think the
staining is not as crisp as using a fresh cut section. Perhaps you can test
this on a positive control slide before using your precious tissue.


Patti Loykasek BS, HTL, QIHC
PhenoPath Laboratories
Seattle, WA




> 
> -----Original Message-----
> From: Bobrowitz, Carol [mailto:carolb <@t> phys.mcw.edu]
> Sent: Friday, April 21, 2006 11:50 AM
> To: Dawson, Glen
> Subject: Help!
> 
> 
> 
> Glen, 
> 
> I would like to send the question below to the histonet but am having
> problems. 
> Could you please post it for me.
> 
> Have a great vacation.
> 
> Thanks, 
> 
> Carol 
> 
> -----Original Message-----
> From:   Bobrowitz, Carol
> Sent:   Friday, April 21, 2006 11:44 AM
> To:     HISTONET (E-mail)
> Subject:        repeating AR heat treatment
> 
> Hello, 
> 
> I have already stained FFPE rat kidney's using FITC with a 2 hour heat
> treatment in citrate buffer ph 6.0.  (the staining was great)
> 
> I am now going to restain the same rat kidney slides with an additional
> primary using DAB as the chromagen.  This primary also requires heat
> treatment.
> 
> Am I correct in my thinking that I should repeat the heat treatment again due
> to the fact that the epitope (binding) sites have already closed after
> accepting the application of the 1st primary?
> 
> Any help will be appreciated.  Thank you in advance.
> 
> Carol Ann Bobrowitz
> Medical College of Wisconsin
> Department of Physiology
> 414-456-8179 
> cbobrowi <@t> mcw.edu 
> 
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