[Histonet] double immuno staining

[Greg Dobbin]

Hi Veronique,
I would suggest you are already on the right track! Perhaps just 
shorten the EIER time, try a different enzyme or try a heat-induced 
epitope retreival method. Good luck.
Greg

From:           	"Veronique Andriessen" <vandries <@t> vub.ac.be>
To:             	"Histonet" <histonet <@t> lists.utsouthwestern.edu>
Date sent:      	Wed, 5 May 2004 16:19:08 +0200
Subject:        	[Histonet] double immuno staining

> Hi folks,
> 
> I need some help.
> 
> I am struggling with a double immunostaining for somatostatin receptor and
> desmin.
> 
> All my previous double stainings worked beautifully, but they were performed
> on fresh frozen acetone fixed pancreatic tissue.
> 
> For staining of somatostatin receptor I have to fix with formaldehyde. I use
> 20 minutes 0.75% PFA  perfused tissue which is snap frozen embedded in
> tissue freezing medium and cut on a cryostat. I was advised to do it this
> way to prevent loss of antigenicity.
> Morphology is quite good.
> This fixation works for the somatostatin receptor staining, but it kills the
> desmin signal.
> I was able to retrieve most of the desmin signal by EIER for 10 minutes with
> trypsin-EDTA, but now the morphology is awful.
> 
> Can someone give me some advice?
> 
> Sincerely yours,
> 
> Veronique Andriessen BAS
> Lab. Molecular Liver Cell Biology,
> Free University Brussels (VUB)Belgium
> 
> 
> 
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~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
Greg Dobbin
Pathology Lab
Atlantic Veterinary College, U.P.E.I.
550 University Ave.
Charlottetown, P.E.I.
Canada,  C1A 4P3
Phone: (902)566-0744
Fax: (902)566-0851
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