Help about BrdU

From: Martha Mesa - Microbiologia (mmesa@javeriana.edu.co)
Date: Tue Apr 10 2001 - 01:15:02 EST


Dear sir,

In my laboratory, I am studying the effect of some compounds on tumoral
cell cycle
by PI staining. In order to know if the antitumoral effect observed is
due to a
decrease in DNA synthesis I am trying to set the BrdU incorporation
assay, by I
have several questions; some of them are:

1. For cells with doubling time of 22  (k562)  and 72 hours (KG1a), how
long must
be the pulse? I have used BrdU 10 uM for 8 hours;  I have only obtained
low
intensity in fluorescence some times.

2.Some protocols recommend more HCl. I have used 2M, 3M and 4M. With 4M
the BrdU signal improves but phase resolution with PI disappear.  I
would like to
know the effect in each phase; how can I obtained that?

Thanks in advance,

Marta Mesa



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