A flow cytometric study of cell death: Failure of some models to correlate with morphological assessment

The balance between cell death and cell proliferation is a significant factor in the growth kinetics of normal and neoplastic tissues. Distinction between the two major forms of cell death, necrosis and apoptosis, is now recognized as important in understanding mechanisms regulating cell survival. A recent approach in the study of apoptosis has been the use of flow cytometry, with some reports indicating that, when stained with propidium iodide (PI), the DNA of apoptotic cells has decreased fluorescence compared with that of viable cells. In this study, we investigated a flow cytometric procedure which used the simultaneous analysis of DNA content and 90 degrees light scatter (90LS). Significant differences in the PI staining pattern and a shift in 90LS were observed when apoptotic death occurred at different stages of the cell cycle. Importantly, such differences only allowed accurate quantification of apoptosis when it occurred in G1. While necrosis could be distinguished from apoptosis when examined during its early stages, a similar staining pattern to that found with apoptosis was observed when necrosis was examined during its latter stages. The results indicate that the measurement of DNA staining cannot be exclusively relied upon to detect apoptosis occurring in all models. However it is useful in the investigation of this process when the death occurs in G1, in that the method offers a rapid means for quantification.