THE USE OF 7-AMINO ACTINOMYCIN-D IN IDENTIFYING APOPTOSIS - SIMPLICITY OF USE AND BROAD-SPECTRUM OF APPLICATION COMPARED WITH OTHER TECHNIQUES

The detection and quantitation of apoptotic cells is becoming increasi ngly important in the investigation of the role of apoptosis in cellul ar proliferation and differentiation. The pathogenesis of hematologic disorders such as aplastic anemia and the development of neoplasia are believed to involve dysregulation of apoptosis. To quantitate accurat ely the proportion of apoptotic cells within different cell types of a heterogeneous cell population such as blood or bone marrow, a method is required that combines the analysis of large numbers of cells with concurrent immunophenotyping of cell surface antigens. In this study, we have evaluated such a method using the fluorescent DNA binding agen t, 7-amino actinomycin D (7AAD), to stain three diverse human cell lin es, induced to undergo apoptosis by three different stimuli, Flow cyto metric analysis defines three populations on the basis of 7AAD fluores cence and forward light scatter, We have shown by cell sorting and sub sequent morphological assessment and terminal deoxynucleotidyl transfe rase-mediated deoxyuridine triphosphate nick end labeling that the pop ulations defined by 7AAD represent live, apoptotic, and late-apoptotic /dead cells. This method is quick, simple, reproducible, and cheap and will be a valuable tool in the investigation of the role of apoptosis in normal physiology and in disease states. (C) 1996 by The American Society of Hematology.