[Histonet] (no subject)

[Favara, Cynthia (NIH/NIAID)]

I have done some of this and will share what I have learned.

Some questions to ask yourself are:

	Do you want to maintain the staining already present?
	Can the second antibody be optimized for the AR that has already
been done?
	Do you know the location of the targets? Different cellular
component, cells, infectious agents, etc?
	

Heat will break the bonds that you have for the first reaction and unless
the reporter is stable - DAB or ALK PHOS RED in my experience- you will lose
any signal that you have currently demonstrated.

In my experience a well fixed and dried specimen  can withstand 2-3 AR with
heat. 

Sometimes it is easier to just recut and stain separately. Depends on what
you are trying to see.

Great resource is Chris Van der Loos' book on multiple staining.


Good Luck,
Cynthia Favara
NIAID/NIH/RML/LPVD
903 South 4th Street
Hamilton, MT 59840
406-363-9317

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-----Original Message-----
From: Bruijntjes, J.P. [mailto:bruyntjes <@t> voeding.tno.nl] 
Sent: Tuesday, November 16, 2004 2:44 AM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] (no subject)

Hi all

 

We have done immunocytochemistry on some slides. An antigen-retrieval
(citrate pH 6.0) was needed. But it would be nice if we could use the same
slides for another antigen (AR-EDTA pH 10 is recommended). Has anyone of you
done this before, and is it possible?  

 

Joost Bruijntjes

Zeist

The Netherlands

 


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