Nuclei isolation for flow analysis

From: Nicholas Terry (ntflow@odin.mdacc.tmc.edu)
Date: Wed Nov 28 2001 - 09:53:49 EST

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At 03:30 PM 11/26/01 +1030, you wrote:

Hi there, I've got another problem for you all...

I'm trying to isolate nuclei from CFSE stained cells so
that I can then fix and permeabilize the nuclei,
perform intranuclear staining, and analyse using flow
cytometry.

<SNIP>
The DNA versus incorporated halogenated pyrimidine (BrdUrd etc) techniques
routinely utilize nuclei. Try  pepsin to digest your cells. The tricks to
avoid clumping include low speed spins and no ++ ions . See our suggested
recipes in  Methods in Cell Biology - Cytometry 3'rd edition or call me for
info.
Nick

Nicholas Terry, M.A., Ph.D.,
Associate Professor,
Experimental Radiation Oncology - 066,
The University of Texas M. D. Anderson Cancer Center,
1515 Holcombe Blvd., Houston, Texas 77030.
'Phone: 713-792-3424,
'Fax: 713-794-5369.
http://drad52.mdacc.tmc.edu.
ntflow@odin.mdacc.tmc.edu.

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