Isolation of highly purified rat cerebral lysosomes using percoll gradients with a variety of calcium concentrations

We were able to isolate a lysosomal fraction from rat brain which had a higher degree of purity than in previous studies with good recovery. This has been made possible by using percoll gradients following the swelling of mitochondria in the presence of calcium which would eliminate contamination from small amounts of mitochondria. By using percoll density gradient centrifugation after the swelling of mitochondria in the presence of calcium, cerebral lysosomes were purified 312-fold with the recovery of approximately 22 %, which is the highest reported for any cerebral lysosomal preparation. The most effective procedure for the separation was achieved by using 1.25 mM calcium incubation with post nuclear fraction. As brain lysosomes may play a major role not only in degrading macromolecules but also in their transport to the deposition site, obtaining purified rat cerebral lysosomes represents an important step in the study of the generation of macromolecules which accumulate in the brain.