A mitochondrial fraction of human platelets that exhibited respiration coupled to ADP utilization has been isolated. These activities were sensitive to classical inhibitors of mitochondrial respiration and oxidative phosphorylation. Pyridine nucleotide-linked substrates were oxidized at low rates (4-7 natoms/min/ mg), but succinate and α-glycerophosphate were oxidized more rapidly (16 and 38 natoms/min/mg, respectively). Difference spectra showed the presence of the mitochondrial respiratory chain cytochromes, c1 + c, b, and a + a3. Electron micrographs revealed the presence of intact mitochondria with well-preserved outer membranes, as well as various α-granules. There appeared to be no qualitative differences between these human platelet mitochondria and pig platelet mitochondria isolated by the same procedure.