In cell culture experiments using bronchoepithelial cells I did the following after overnight incubation at 37 C 5% CO2. 1) Aspirate and save cells. 2) Add trysin to all wells for 5 minutes. (I did preliminary exps to show that trypsin did not increase the uptake of annexin. By the way I never could get EDTA to cause cells to detact.) 3) Neutralize trypin with FCS containing media 4) Do AV protocol. On Fri, 26 Feb 1999, ronald hill wrote: > > Has anyone used annexin V staining to detect apoptosis in rat > epithelial cells. We are having trouble obtaining intact cells after > staining. A majority of the cells become PI positive after removal from > the plate. Any tricks? Should we remove cells first and then stain? > > Thanks for any suggestions. > > -Ron Hill > > Ronald J. Hill, Ph.D. > Group Leader, Immunology Targets > Sugen, Inc. > 230 East Grand Avenue > South San Fransisco, CA 94080-4811 > > Tel: (650) 553-8797 > Fax: (650) 553-8313 > E-mail: ron-hill@sugen.com > > >
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