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TUNEL/PI staining inconsistency - (Sep/26/2008 )

Hi to everybody !!!!
In order to quantify apoptotic cell death (by flow cytometry) I noticed that in my model (cell lines treated with Sturosporin) there is a big difference between data coming from TUNEL staining and canonical PI staining, in terms of % of apoptotic cells !?!?!?! In detail, using the PI staining protocol (cells fixed with MetOH-Acetone buffer) I can see positive cells that are negative to TUNEL staining (BD kit) !?!?!?!?!
I'm sure this are not necrotic as they are able to exclude PI when not fixed (even if necrotic cells should be also TUNEL positive) !!!!

Any explanation or suggestion ??????


From my understanding, TUNEL staining only stain cells that are undergoing apoptosis. PI staining without fixing, both necrotic and apoptotic cells will be stained. For apoptotic cells, by flow cytometry, apoptotic cells will have positive TUNEL and PI stainng. If only PI positive, those will be necrotic cells.