[Histonet] nuclear counterstain

[Favara, Cynthia (NIH/NIAID)]

Atoska,

I have used the Methylgreen nuclear counter stain from Chris van der Loos
with VIP. It is a s follows:

Methylgreen, Crystal Violet-free [Sigma M6776]
0.1% methylgreen in sodium acetate buffer [ 100mM, pH 5.2]
Solution stable @4C for 1 month

Cover section with sodium acetate buffer [10mM, pH5.2]  15 min
Blot off do not wash
Cover with methyl green solution 5 min
Wash briefly with distilled water
Dehydrate and mount organically


This is out of his book Immunoenzyme Multiple Staining Methods

Fabulous resource

c



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


-----Original Message-----
From: Atoska S. Gentry [mailto:gentras <@t> vetmed.auburn.edu]
Sent: Friday, September 12, 2003 9:15 AM
To: Histonet
Subject: [Histonet] nuclear counterstain



Hello again, guess I need to rephrase my initial inquiry on the 
counterstain to use with the Vectastain ABC-HRP kit. But, first special 
thanks to Dr. John Kiernan. He always replies with such informative and 
thorough remarks. Please pardon my partial entry of facts. This inquiry is 
on behalf of a couple of colleagues of mine. And yes their work was with 
monolayers of cultured bone marrow cells. However, the HRP was detected 
with the Vector VIP substrate       ( for which Vector Labs does not reveal 
the contents) and yielded neurons that are purple in color. Therefore, we 
need something like acridine orange which stains DNA. Has anyone had any 
experience/success using acridine orange or something similar with this 
system that yields the same results? Thanks, Atoska


Atoska S. Gentry B.S., HT(ASCP)
Research Assistant III
Scott-Ritchey Research Center
College of Veterinary Medicine
Auburn University, AL  36849
Phone# (334)844-5579  Fax# (334)844-5850 


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