OBJECTIVETo establish a method for simultaneous isolation and primary culture of rat hepatocytes, hepatic stellate cells, Kupffer's cells and hepatic sinus endothelial cells.

METHODSBy combining in situ perfusion, in vitro perfusion, density gradient centrifugation and differential adhesion, primary rat hepatocytes, hepatic stellate cells, Kupffer's cells and hepatic sinus endothelial cells were obtained. The purity of these cells were assessed with morphological observation, immunofluorescent staining and ink phagocytosis assay.

RESULTSWe successfully obtained the 4 primary cells simultaneously by combining in situ perfusion with in vitro perfusion, density gradient centrifugation, and differential attachment. The cell yield rate, cell viability and purity all met requirements for the subsequent cell experiment.

CONCLUSIONThe combined cell isolation and culture method is feasible to isolate primary rat hepatocytes, hepatic stellate cells, Kupffer's cells and hepatic sinus endothelial cells simultaneously.