[Histonet] Re: alph-MSH

[Kate Ellacott]

Hi Leigh,

The one that we tend to use is the sheep anti-alpha MSH from Chemicom,
AB5087. Is this the one that you have tried? We use it at 1:6000 on
free-floating perfused brain tissue. In my experience it only works in
the hypothalamus (ARC) but not in the brainstem (if you know of one that
works in the brainstem let me know!). From your message it sounds like
you are getting a signal just with a lot of background. Have you tried
increasing your blocking serum concentration? I routinely use 5% donkey
serum to block for 2h at room temp and then incubate in primary antibody
diluted in blocking serum overnight at 4 degrees C. 

What secondary are you using? You don't say if you are using HRP or
fluoresence. Jackson Immunoresearch have a number of secondary
antibodies that have been preabsorbed to minimize cross reactivity and
background. These are good.

Please feel free to e-mail me if you have any more questions as I know
how frustrating this can be.

Best wishes,

Kate

Dr. Kate Ellacott
Vollum Institute
Portland, OR
USA


Message: 1
Date: Thu, 15 Apr 2004 07:43:54 -0500
From: Robbie L Townsend <townserl <@t> lsu.edu>
Subject: [Histonet] alphaMSH
To: histonet <@t> lists.utsouthwestern.edu
Message-ID:
	<OFA02A33E6.8E68C156-ON86256E77.0045F01E-86256E77.0045F028 <@t> lsu.edu>
Content-Type: text/plain; charset=US-ASCII





Hi histonetters.  Does anyone out there know of a good alpha-MSH
antibody
to use on mouse brain and the appropriate dilution?  I have tried one
from
Chemicon and one from Immunostar, but I am still getting very high
background.  I am using them at a very high dilution, rinse very well,
and
block for two hours.  Any help would be greatly appreciated.  Thanks

Leigh Townsend
Research Associate
Pennington Biomedical
Baton Rouge, La
email- townserl <@t> lsu.edu
phone- (225)763-3001




Dr. Kate Ellacott
Vollum Institute
Oregon Health and Science University
3181 SW Sam Jackson Park Rd
Portland
OR 97239

(503) 494 4667