Staining with PI & BrdU

Thomas Delohery t-delohery at ski.mskcc.org
Thu Mar 28 01:55:30 EST 1996


Hi,

>I've been staining cells with BrdU & getting about 60-80% incorporation
>after a 2h pulse - is this the norm?  The cycling time of these cells is 12
>h.  I've tried adding PI to look at DNA content & find that 48% of the cells
>are in S phase.  Do these results make sense?  I'm not a DNA person so I'm
>not quite sure.  Someone that I spoke to thought that it would make sense,
>but my supervisor doesn't think so.  So, I thought I'd ask the opinion of
>people who do this everyday!

 Makes perfectly good sense to me.  Are thes MEL cells?  CHO? ~50% S-phase
with a 12 hr. doubling time  tells you these are relatively rapid growers.
Keep in mind the BrdU is labelling ALL cells in S over a 2hr. period - cells
in late S at the beginning of 2h and cells entering S 2hrs later.
This, as opposed to a the % S measured by PI at the instant of fixation.

good luck,
td

--
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 Thomas Delohery                        | Internet: t-delohery at ski.mskcc.org
 Manager, Flow Cytometry Core Facility  |    Phone: (212) 639-8729
 Memorial Sloan-Kettering Cancer Center |      Fax: (212) 794-4019
 1275 York Ave. Box 98                  |
 New York, NY    10021                  |
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