[Histonet] Re: fine red ppt on sections after AP Envision.

[Gayle Callis]

In addition to Joanne's reply, using PBS rinsing at chromogen steps can 
provide phosphate ions that will react with the AP.  Either use TRIS 
buffered saline throughout whole staining protocol or at least do a TBS 
rinse x 3 before the AP chromogen step to get rid of phosphate ions.

Another nice block for endogenous Alk phos is KPL's ready to use universal 
blocker, done at beginning of staining protocol ( just like peroxidase 
blocking, up front).    Nice thing is you don't have to add anything to the 
chromogen at the end.   This block nicely replaces levamisole.



At 09:10 AM 11/18/2004, you wrote:
>Dr. Montgomery,
>Have you tried adding levamisole to your chromogen? It blocks endogenous 
>alk-phos in tissue.Dako sells it seperately from kit. I use 1 drop per ml. 
>of chromogen, and it works quite well.
>Jo
>
> >>> "Ian Montgomery" <ian.montgomery <@t> bio.gla.ac.uk> 11/18/04 10:52AM >>>
>          I wonder if anyone can offer any hints and tips for Dako Envision.
>Localisation is fine, HRP is clean but alkaline phosphatase, despite my
>efforts, is still giving me quite a lot of background. This manifests
>itself as a fine red ppt. over the specimen.
>Ian.
>
>Dr. Ian Montgomery,
>Histotechnology,
>Graham Kerr Building,
>Institute of Biomedical & Life Sciences,
>University of Glasgow,
>Glasgow,
>G12 8QQ.
>Tel: 0141 339 8855
>Office: 4652
>Lab: 6644.
>Pager: 07625 702883
>e-mail: ian.montgomery <@t> bio.gla.ac.uk
>
>
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Gayle Callis
MT,HT,HTL(ASCP)
Research Histopathology Supervisor
Veterinary Molecular Biology
Montana State University - Bozeman
PO Box 173610
Bozeman MT 59717-3610
406 994-6367 (lab with voice mail)
406 994-4303 (FAX)