Distribution of peroxidase activity was investigated electron cytochemically by the diaminobenzidine (DAB) method in the fixed cells of Tetrahymena pyriformis strain W. Peroxidase with predominantly high activity was located in the mitochondria as shown by electron-opaque deposits of reacted DAB, but not in other cytoplasmic organelles. DAB products were deposited on mitochondrial membranes: outer membranes, inner membranes and cristate membranes. Ultrastructural sites of peroxidase activity were clearly identified on the outer sides of mitochondrial cristate membrane structures facing the intracristal spaces. Significantly, electron-dense products of DAB were also visualized on the mitochondrial inner membranes and the outer membranes. However, finer locations of peroxidase in these membrane structures were less clear. The enzyme was inactivated by heating for 10 min at 50°C or by treatment with methanol-nitroferricyanide, an inhibitor of peroxidase. Inhibition by 1 mM potassium cyanide was fairly intensive but not complete. Ten millimolar 3-amino-l,2,4-triazole, a catalase inhibitor, and 10 mM sodium azide (from neutral pH through alkaline pH) had no effects. Sodium azide, however, suppressed all peroxidase activity at pH 5. These enzymatic characteristics are similar to those of cytochrome peroxidases in the mitochondria of baker's yeast and in the cells of Thiobacillus novellus or of Pseudomonas fluorescens.