M phase cells
ZBIGNIEW DARZYNKIEWICZ (darzynk@nymc.edu)
Sat, 13 Jul 1996 14:49:06 -0500
Dr. Hill inquired about the methods alternative to AO which can be
used to distinguish M from G2 phase cells by flow cytometry.
Because apoptotic cells, as mitotic, have DNA very sensitive to
denaturation, they may be mistakenly classified as the latter (and
vice versa). Hence, the AO method may not work well when there is a
significant number of cells dying by apoptosis in the measured
population.
A very reproducible, alternative to AO, method of discrimination of
mitotic from G2 cells is based on bivariate analysis of cyclin A and
DNA content (Gong et al., Exp. Cell Res. 220, 226-231, 1995). In
contrast to G2 cells which express cyclin A, mitotic cells are cyclin
A negative. This cyclin is rapidly degraded at the stage of
prometaphase. Tha cyclin A antibodies which are applicable to
immunocytochemical detection of this protein are commercially
available (e.g. from PharMingen, clone BF683). Besides, cyclin A,
unlike other cyclins, was expressed in a very "scheduled" pattern in
all the cell lines (> 20) that we have studied.
Zbigniew Darzynkiewicz
CD ROM Vol 2 was produced by staff at the Purdue University Cytometry Laboratories
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If you have any comments please direct them to Dr. J. Paul Robinson, Professor & Director, PUCL,
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