Separation of rat liver lysosomes and mitochondria in the A-XII zonal centrifuge

Rat liver lysosomes and mitochondria were isolated free of mutual cross-contamination on the basis of differences is sedimentation velocity during a single run in the type A-XII zonal centrifuge. Cytochrome oxidase was employed as the biochemical marker for the mitochondria, and acid phosphatase (nitrophenyl phosphate substrate, sodium acetate buffer, pH 5.0, in the presence of the non-ionic detergent Turgitol NPX) was employed as the biochemical marker for the lysosomes. Structure-linked latency and activation was demonstrated for the "lysosomal" phosphatase following exposure to Turgitol NPX, osmotic lysis by treatment with distilled water, and fragmentation in a Waring blender. Electron microscopy of the isolated fractions showed that the microsomes were contaminated by glycogen but free of mitochondria and lysosomes, the lysosomes were contaminated by microsomes and glycogen but were free of mitochondria, and the mitochondria were cross-contaminated only by a few microsomal vesicles.