second reagent choice

From: Howard Ratech (ratech@aecom.yu.edu)
Date: Tue Apr 20 1999 - 14:08:24 EST


Hi flowers:

I would appreciate it if you could give me some advice on the following
sort/analysis problem. Analysis will be done on a FACscan with one laser.

We are staining human tumor nuclei isolated from paraffin blocks (a la
Hedley):

1.	primary unlabelled rabbit anti-human p53
	secondary FITC-conjugated swine anti-rabbit

2. 	propidium iodide (PI; for nuclear DNA content)


Combination 1 plus 2 has been successful. Now we also want to
simultaneously measure BCL6.

3.	mouse monoclonal anti-human BCL6, unlabelled
	(anti-BCL6 is not commercially biotinylated or labelled,
	to my knowledge)
	What fluorochrome on what polyclonal species antisera do you
	recommend?

	Or alternatively, has anyone actually used FITC, PE, 7AAD to
detect 2 protein molecules and DNA content/ploidy? Is 7AAD inferior to PI
for nuclear DNA content/ploidy? 

Thanks.

Howard

Howard Ratech, M.D.
Albert Einstein College of Medicine/
Montefiore Medical Center
111 E 210th St
Bronx, NY 10467



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