Spontaneous and Fas/APO-1–mediated apoptosis of freshly isolated human neutrophils. (A) Cytospin preparations of neutrophils were stained with hematoxylin/eosin and apoptotic cells were quantified morphologically: (1) medium alone 0 hour, (2) medium alone 6 hours, (3) anti-Fas MoAb 250 ng/mL 6 hours, (4) medium alone 24 hours. Bold arrows indicate typical cells in the intermediate stages of apoptosis. Percentages of apoptotic cells are reported in Table 1. (B) Agarose gel electrophoresis of DNA showing a time-dependent increase in the formation of oligonucleosomal DNA fragments in spontaneous (lane 1, 0 hour; lane 2, 3 hours; lane 3, 6 hours; lane 4, 12 hours; lane 5, 24 hours) and Fas/APO-1–triggered (lane 6, 3 hours; lane 7, 6 hours; lane 8, 12 hours) apoptosis. Lane M represents molecular weight marker (kB). (C) Flow cytometric analysis of annexin V binding in neutrophils undergoing spontaneous (panel 1, 6 hours; panel 2, 24 hours) and Fas/APO-1–mediated (panel 3, 6 hours; panel 4, 24 hours) apoptosis. The percentage of PS positive cells in each sample is indicated. Similar results were obtained in four separate donors (see Table 1).