Apoptosis & surface markers

Neal Benson (nbenson@nervm.nerdc.ufl.edu)
Fri, 2 Jun 95 18:36:58 edt

Regarding the recent discussions regarding methods of identifying surface
antigens and apoptosis, I thought this may be of interest. We presented a
poster at the recent Am. Soc. for Clin. Oncology Meeting in Los Angeles, an
adaptation of the so called "tunel" assay to include surface antigen
staining. We stain first for the surface antigens using direct
PE-conjugates, treat the cells with Facslyse (sold by BD), then apply the
tunel method using biotin-dUTP and FITC-streptavidin. Facslyse seems to
sufficiently permeablize the cells, yet keep the surface staining and the
cut DNA stable (though it's not suitable for cell cycle due to broad CV's).
We've applied it to KG1 cells (which express CD34), as well as a few other
culture lines.

When we had tried alcohol fixation, we found a reduction in CD34 antibody
fluorescence, as has been observed by others.

A manuscript is in the works. I'd be happy to send out a copy of the
"methods" section of the poster to anyone interested.

Neal
=======================================================================
Neal A. Benson Tel: (904) 392-0008
University of Florida Fax: (904) 392-4693
Department of Pathology Email: nbenson@nervm.nerdc.ufl.edu
Box 100275
Gainesville, FL 32610-0275 USA

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