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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