A new method (type B) for isolation of canine islets was developed, utilizing intraductal collagenase perfusion, stationary digestion, filtration and tissue separation by means of a dextran density gradient. The results of this technique were compared with those of a previously established method (type A). Islets were autotransplanted either via the splenic or the portal vein. Lasting normoglycemia was obtained in 5/8 intrasplenic type B transplants (63%), 6/8 intrahepatic type B transplants (75%), 5/6 intrasplenic type A transplants (83%) and 0/3 intrahepatic type A transplants. No difference was found in metabolic parameters 2 weeks after successful transplantation following type A or B islet isolation. Islet yield was higher with type A and purification better with type B isolation. Both techniques are relatively simple and inexpensive. Because of its higher purification rate and the success of intrahepatic islet transplantation method B has replaced method A as the routine procedure in our laboratory.