> Martin Kelly wrote:
> >
> > > we would like to ask some advice r.e. the interpretation of some
> > work we have been doing on induced sputum and BAL from normal
> > subjects and COPD patients. We have been using 'flow' to look at
> > the granulocytes, and examine the AV and PI staining in the
> > events which we have gated on, which we feel are in the region
> > in which one would expect to find granulocytes. Cells coming up
> > AV+PI- we interpret as apoptotic; AV+PI+ as necrotic.
>
Bear in mind that late stage apoptotic cells will no longer
exclude
PI. The best identification for the process of aopoptosis is
fluorescence microscopy after labeling with a DNA stain [PI, DAPI, AO,
etc.]. You should be able to identify those cells undergoing apoptosis
and determine if "necrosis" is actually present in your system.
Alternately,run a 2 nd assay such as TUNEL or subdiploid assay to
verify
your annexin results.
Regards,
Tom
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Thomas W. Mc Closkey, Ph. D., Director of Flow Cytometry
North Shore University Hospital, New York University School of Medicine
Boas Marks Biomedical Research Center, 350 Community Drive
Manhasset, Long Island, New York 11030
ph: 516-562-4844 [office], 516-562-1135/4641 [lab] fax: 516-562-2866
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