Dr. Schmidt asks for explanation why the results with annexin V do not correlate with TUNEL. The problem is that so called "atypical apoptosis", i.e. cell death when one or another of supposedly classical apoptotic features is missing, is quite common. Internucleosomal DNA fragmentation that is detected by the TUNEL reaction may occur or not and there are dozens of publications (different cell types, different inducers of apoptosis) describing apoptosis when DNA fragmentation stopped at 50-300 kb sections and have not progressed to the internucleosomal DNA sections. Such fragmentation is not easily detected by the TUNEL method. Likewise there are situations when cells bind annexin V but the binding has no relation to apoptosis. Morphological changes (cell shrinkage, chromatin condensation- reflected by DNA hyperchromicity) still remain the "gold standard" to identify apoptotic cells. These and other problems and pittfals in detection of apoptosis were described in Methods in Cell Biology, Vol. 63, 527-544, 2001. Numerous authors define now apoptosis as the "caspases-mediated cell death". We are very satisfied using the FLICA methodology to detect caspases activation (Smolewski et al. Cytometry, 44: 72-82, 2001). Zbigniew Darzynkiewicz Brander Cancer Research Institute New York Medical College 19 Bradhurst Ave. Hawthorne, NY 10532 tel: 914-347-2801 fax: 914-347-2804 http://www.geocities.com/z_darzynkiewicz
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