NADH-cytochrome b5 reductase of rat liver microsomes was efficiently solubilized by incubation with the lysosomal fraction of the same tissue. Examination of optimum conditions for the solubilization suggested the participation of a lysosomal enzyme (or enzymes) in the release of the reductase from microsomal particles. Available evidence denied the principal role of Naja naja venom in the solubilization of the reductase reported by previous workers.Solubilization of microsomal enzymes by lysosomal digestion showed a peculiar specificity. While most of NADH-cytochrome b5 reductase [EC 1.6.2.2] was easily made soluble by the treatment, NADPH-cytochrome c reductase and cytochrome b5 were solubilized to much lesser extents. Only a small portion of microsomal protein was solubilized, and phospholipid was not released detectably from microsomes.In contrast to lysosomes, digestion with trypsin [EC 3.4.4.4], Nagarse, and other proteinases failed to solubilize NADH-cytochrome b5 reductase, whereas these proteinases easily solubilized NADPH-cytochrome c reductase and cytochrome b5 from microsomes. The nature of the effective solubilizing factor in lysosomes is still to be elucidated.NADH-cytochrome b5 reductase activity was considerably stimulated when the reductase was solubilized from microsomes by lysosomal digestion. NADH-ferricyanide reductase activity showed no stimulation by the treatment. However, these two activities were released in parallel from microsomes, and seem to be associated with the same enzyme. NADH-cytochrome b5 reductase of rat liver microsomes was efficiently solubilized by incubation with the lysosomal fraction of the same tissue. Examination of optimum conditions for the solubilization suggested the participation of a lysosomal enzyme (or enzymes) in the release of the reductase from microsomal particles. Available evidence denied the principal role of Naja naja venom in the solubilization of the reductase reported by previous workers. Solubilization of microsomal enzymes by lysosomal digestion showed a peculiar specificity. While most of NADH-cytochrome b5 reductase [EC 1.6.2.2] was easily made soluble by the treatment, NADPH-cytochrome c reductase and cytochrome b5 were solubilized to much lesser extents. Only a small portion of microsomal protein was solubilized, and phospholipid was not released detectably from microsomes. In contrast to lysosomes, digestion with trypsin [EC 3.4.4.4], Nagarse, and other proteinases failed to solubilize NADH-cytochrome b5 reductase, whereas these proteinases easily solubilized NADPH-cytochrome c reductase and cytochrome b5 from microsomes. The nature of the effective solubilizing factor in lysosomes is still to be elucidated. NADH-cytochrome b5 reductase activity was considerably stimulated when the reductase was solubilized from microsomes by lysosomal digestion. NADH-ferricyanide reductase activity showed no stimulation by the treatment. However, these two activities were released in parallel from microsomes, and seem to be associated with the same enzyme.