A Novel Efficient Technique of Pancreatic Islet Cell Isolation and Purification

Objective:To explore the method of obtaining enough mouse islets with high purity in order to set up an animal model for clinical islet transplantation.Methods: The 6~8 weeks male C57BL/6 mice weighing 25~30 g were intraperitoneal anesthetized before the operation of common bile duct ligation.Then we isolated and purified mice islets by adopting collagenⅤ retro perfusion,situ digestion,gradient centrifugation in Ficoll-400 solution and sorted islets using sterile capillary pipettes in the phase contrast microscope.Islets purity was assessed by dithizone staining.Eventually,we cultured islet cells and observed the shape of islets on day 3,8 and 24 separately.Results: From this way,we could obtain 390±20 islets in each mouse.The isolated islets were round or mass,50-150 μm in diameter,complete and bright.The purity of the islets was more than 85%.Conclusions: We improve the method of isolating islets including retro perfusion,situ digestion and gradient centrifugation in Ficoll-400 solution,which is timesaving and effective.The cultured islet cells had high activity and formed single cells in petri dishes within 24 days,which is suitable for further study.