胰蛋白酶和胶原酶灌注法分离提取小鼠原代肝细胞的比较  

Comparison of isolation and extraction of primary mouse hepatocytes by trypsin and collagenase perfusion

作　　者：任文洁 林哲绚 REN Wenjie;LIN Zhexuan(Bioanalytical Laboratory,Shantou University Medical College,Shantou 515041,China)

机构地区：[1]汕头大学医学院生物分析实验室,广东汕头515041

出　　处：《汕头大学医学院学报》2022年第4期204-209,共6页Journal of Shantou University Medical College

基　　金：广东省科技专项资金(210705156884432);广东省自然科学基金(2018A030307005)。

摘　　要：目的:胰蛋白酶和胶原酶灌注法分离提取小鼠原代肝细胞的比较。方法:改良Seglen两步灌注法,选用胰蛋白酶和胶原酶D分别对同批生长的10~16周C57BL/6J雄性小鼠进行在体灌注分离肝实质细胞,比较分离肝细胞的存活率、纯度、形态、活性、获取数量等。结果:两种方法分离得到的小鼠原代肝细胞的存活率和纯度均达到90%以上,且在形态学上无明显差异,不同培养时间的细胞活性基本一致。胶原酶灌注法所得到的细胞数量多于胰蛋白酶灌注法。结论:胰蛋白酶和胶原酶灌注法均适用于分离提取小鼠原代肝细胞,可根据实验条件选择不同的灌注方法。Objective:To compare the separation and extraction of mouse primary hepatocytes by trypsin and collagenase perfusion methods.Methods:A modified Seglen two-step perfusion method was used to isolate hepatic parenchymal cells by in vivo perfusion of trypsin and collagenase D in male C57BL/6J mice grown in the same batch from 10 to 16 weeks,respectively,to compare the survival rate,purity,morphology,viability,and number of isolated hepatocytes obtained.Results:The survival rate and purity of mouse primary hepatocytes isolated by the two methods were both above 90%,and there was no significant difference in morphology.The cell viability was basically the same at different culture times.The number of cells obtained by the collagenase perfusion method was significantly better than that obtained by the trypsin perfusion method.Conclusion:Both trypsin and collagenase perfusion methods are suitable for the isolation and extraction of mouse primary hepatocytes,and different perfusion methods can be selected according to the experimental conditions.

关 键 词：酶灌注法 小鼠原代肝细胞 胰蛋白酶 胶原酶 

分 类 号：R-332[医药卫生]