Hi Ellen... More and more we've been doing early-stage apoptotic analysis on our laser scanning cytometer (LSC) to avoid the cell trauma associated with detachment. Although later apoptotic cells do tend to round up and detach from their growth surface (thus eliminating themselves from the assay), we can still find many earlier apoptotic cells on the LSC using annexin V, caspase substrates, mito membrane potential or more permeable DNA dyes such as 7-AAD. Annexin V binding is not the only apoptosis assay adversely affected by trypsinization - we often get ambiguous results from our other apoptotic assays as well after cell removal, since trypsinization can alter membrane permability, mito membrane potential, etc. In general our LSC data is much cleaner. Bill Telford ETIB-CCR-NCI-NIH on At 10:08 PM 8/1/01 +0200, Ellen.Freed@astrazeneca.com wrote: > >Since we're on this subject. I've been told it's an early marker. . . >I have a separate question. Is anyone doing Annexin V staining on adherent >cells. I understand that once you trypsinize the cells, this is no longer >an accurate measure. Any comments on this or ways to get around it? > >Thanks > >Ellen Freed > >Ellen Freed >Scientist >AstraZeneca R&D Boston
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