Caspase-3–like activity in neutrophils undergoing spontaneous and Fas/APO-1–mediated apoptosis, as measured by cleavage of the specific fluorogenic substrate DEVD-AMC (50 μmol/L). (A) Time course of caspase-3–like activation in spontaneous (⧫) and anti-Fas MoAb-triggered (250 ng/mL; ◊) apoptosis of neutrophils. The cells were lysed at various times (0 to 72 hours) after initiation of in vitro culture and the release of AMC was monitored. The maximum rate of AMC release (pmol/min) was estimated by linear regression (r² > 0.99). Data shown are the mean and range of duplicate determinations from a representative experiment. (B) The competitive inhibitor DEVD-CHO (50 nmol/L) was added to cell lysates in vitro and shown to block DEVD-AMC cleavage in cells undergoing spontaneous apoptosis (control, ○; plus inhibitor, •). Cells (1 × 10⁶) were incubated for 15 hours before the fluorometric assay. Each data point represents the average value of determinations performed in triplicate samples.