>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
-- ============================================================================== Thomas Delohery | Internet: t-delohery@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 | ==============================================================================
CD-ROM Vol 3 was produced by Monica M. Shively and other staff at the
Purdue University Cytometry Laboratories and distributed free of charge
as an educational service to the cytometry community.
If you have any comments please direct them to
Dr. J. Paul Robinson, Professor & Director,
PUCL, Purdue University, West Lafayette, IN 47907.
Phone: (765)-494-0757;
FAX(765) 494-0517;
Web