原代大鼠肝细胞的分离和培养  

Isolation and Culture of Primary Rats Hepatocytes

作　　者：蒋永生[1] 程向东[2] 刘文洪[1] 项海[1] 姚立[1] 

机构地区：[1]浙江中医药大学,浙江杭州310053 [2]浙江省肿瘤医院,浙江杭州310022

出　　处：《中华中医药学刊》2013年第5期1111-1112,I0008,共3页Chinese Archives of Traditional Chinese Medicine

基　　金：浙江省钱江人才计划资助项目(Q1770802006);浙江省自然科学基金资助项目(Y2101290)

摘　　要：目的:探讨原代大鼠肝细胞分离及培养的简易方法,建立稳定简便的大鼠原代肝细胞分离培养方法。方法:采用改良原位两步循环灌流法及低速离心法分离原代大鼠肝细胞,常规DMEM:高糖培养基培养原代大鼠肝细胞,测定1周内生长情况,倒置显微镜及扫描电镜观察肝细胞形态变化。结果:倒置显微镜下观察肝细胞在接种后3 h基本完成贴壁,贴壁的细胞呈卵圆形,24 h后胞体变大变平,随后细胞相互靠拢呈岛状或条索状连接,3 d后扫描电镜下细胞呈现良好形态。结论:用改良原位两步循环灌注法及低速离心法成功分离并培养原代大鼠肝细胞,可操作性强,可在具备基本细胞培养条件的实验室推广。Objective：To investigate a simple and feasible method for primary rats hepatocytes isolation and culture,and to establish a stable and easy methods of isolation and culture of primary rats hepatocytes.Methods：Hepatocytes were isolated by using a modified two-step in-situ-circulating perfusion method followed by low-speed centrifugation.The hepatocytes were cultured in Dulbecco＇s modified Eagle＇s medium with high level of glucose.The viabilities of cells were evaluated within one week.Cellular morphological changes were observed by using an inverted microscope and scanning electron microscope（SEM）.Results：The elementary adherence completion of the hepatic cells had been posted inoculation after 3h under inverted microscope,the cells of adherence offerred ovate.The hepatocytes enlarged and flattened round,attached to the surface of collagen-coated dishes 24h after seeding and then connected with surrounding cells,forming islands or trabs.After 3 days,cells were well under the SEM.Conclusions：Primary rats hepatocytes can be successfully isolated and cultured using a modified two-step in-situ-circulating perfusion method,which is highly feasible.This method is available to perform in labs with basic cell culture conditions.

关 键 词：大鼠 原代肝细胞 分离 培养 

分 类 号：R-332[医药卫生]