AIM:To explore the methods of hepatocytes culture in acollagen gel mixture or between double layers of collagensandwich configuration and to examine the functional andcytomorphological characteristics of cultured hepatocytes.METHODS:A two-step collagenase perfusion techniquewas used to isolate the hepatocytes from Wistar rats ornewborn Chinese experimental piglets.The isolatedhepatocytes were cultured in a collagen gel mixture orbetween double layers of collagen sandwich configurationrespectively.The former was that rat hepatocytes weremixed with type I rat tail collagen solution till gelled,andthe medium was added onto the gel.The latter was thatswine hepatocytes were seeded on a plate precoated withcollagen gel for 24h,then another layer of collagen gelwas overlaid,resulting in a sandwich configuration.Thecytomorphological characteristics,albumin secretion,andLDH-release of the hepatocytes cultured in these twomodels were examined.RESULTS:Freshly isolated rat hepatocytes were successfullymixed and fixed in collagen gel,and cultured in the gelcondition.During the culture period,the urea synthesizedand secreted by rat hepatocytes was detected throughoutthe period.Likewise,newborn experimental piglethepatocytes were successfully fixed between the doublelayers of collagen gel,forming a sandwich configuration.Within a week of culture,the albumin secreted by swinehepatocytes was detected by SDS/PAGE analysis.The typicalcytomorphological characteristics of the hepatocytes culturedby the above two culture models were found under a phase-contrast microscope.There was little LDH-release duringthe culture period.CONCLUSION:Both collagen gel mixture and double layersof collagen sandwich configuration can provide culturalconditions much closer to in vivoenvironment,and are helpfulfor maintaining specific hepatic functions and cytomorphologicalcharacteristics.A collagen gel mixture culture may be moreeligible for the study of bioartificial livers. AIM: To explore the methods of hepatocytes culture in acollagen gel mixture or between double layers of collagens andwich configuration and to examine the functional and cytomorphological characteristics of cultured hepatocytes. METHODS: A two-step collagenase perfusion technique was used to isolate the hepatocytes from Wistar rats or newborn Chinese experimental piglets. The isolated hepatocytes were cultured in a collagen gel mixture or between double layers of collagen sandwich configurationrespectively. The former was that rat hepatocytes were fixed with type I rat tail collagen solution till gelled, and the medium was added onto the gel. latter was thatswine hepatocytes were seeded on a plate precoated with collagen gel for 24h, then another layer of collagen gel was overlaid, resulting in a sandwich configuration. The cytomorphological characteristics, albumin secretion, and LDH-release of the hepatocytes cultured in these twomodels were observed .RESULTS: Freshly isolated rat hepatocytes were success fullymixed and fixed in collagen gel, and cultured in the gelcondition. During the culture period, the urea synthesized and secreted by rat hepatocytes was detected throughout the period. Liverwise, newborn experimental piglethepatocytes were successfully fixed between the doublelayers of collagen gel, forming a sandwich configuration. Within a week of culture, the albumin secreted by swinehepatocytes was detected by SDS / PAGE analysis. Typical cytomorphological characteristics of the hepatocytes cultured by the above two culture models were found under a phase-contrast microscope. There was little LDH-release during the culture period. CONCLUSION: Both collagen gel mixture and double layers of collagen sandwich configuration can provide cultural conditions are much closer to in vivoenvironment, and are helpful for maintaining specific hepatic functions and cytomorphological characteristics. A collagen gel mixture culture may be more eligible for the study of bioartificial livers.