[Histonet] tricks about antibody generated in human on human tissue for IHC

[Connolly, Brett M]

Hi Carole,

While I have no direct experience with human on human IHC, my first thought would be to biotinylate your primary antibody and then you would omit the anti-human secondary which is the most likely source of your background staining. 

If you are using fluorescence you could also conjugate your primary with a fluorophore  (http://www.invitrogen.com/site/us/en/home/brands/Molecular-Probes/Key-Molecular-Probes-Products/Zenon-Labeling-Technology.html)



Brett M. Connolly, Ph.D.
Principal Scientist, Imaging Dept.
Merck & Co., Inc.
PO Box 4, WP-44K
West Point, PA 19486
brett_connolly <@t> merck.com
T- 215-652-2501
F- 215-993-6803





-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of carole.chotard <@t> fr.netgrs.com
Sent: Friday, January 11, 2013 5:32 AM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] tricks about antibody generated in human on human tissue for IHC

Dear all,

 

I work in an histopathology department and we intend to do
immunohistochemistry with an antibody generated in human on human
tissues. So far, we don't have any experience with that. So I was
wondering if somebody would have some feedback to prevent strong
background generated by this human antibody on human tissue. We are
planning in a first place to decrease the concentration of the secondary
antibody Donkey <@t> human which is recommended at 1/200 by Jackson, using it
at 1/500 for example.

 

But if anybody would have some experience with this kind of experiments,
their tricks would be more than welcome.

 

Best Regards,

 

Carole Chotard

 

 

Carole Chotard, Ph.D.

Pharmacology Research Scientist

Molecular Pharmacology and Pathophysiology Department

IDRS

125 Chemin de Ronde

78290 Croissy-sur-Seine

France

Phone : +33 1 55 72 29 48

Fax :       +33 1 55 72 25 96

carole.chotard <@t> fr.netgrs.com

 

 

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