Mitochondria are known to combine life-supporting functions with participation in apoptosis by controlling caspase activity. Here, we report that in human blood neutrophils the mitochondria are different, because they preserve mainly death-mediating abilities. Neutrophil mitochondria hardly participate in ATP synthesis, and have a very low activity of the tested marker enzymes. The presence of mitochondria in neutrophils was confirmed by quantification of mitochondrial DNA copy number, by detection of mitochondrial porin, and by JC-1 measurement of Deltapsi(m). During neutrophilic differentiation, HL-60 cells demonstrated a profound cytochrome c depletion and mitochondrial shape change reminiscent of neutrophils. However, blood neutrophils containing extremely low amounts of cytochrome c displayed strong caspase-9 activation during apoptosis, which was also observed in apoptotic neutrophil-derived cytoplasts lacking any detectable cytochrome c. We suggest that other proapoptotic factors such as Smac/DIABLO and HtrA2/Omi, which are massively released from the mitochondria, have an important role in neutrophil apoptosis.