SD乳鼠心肌细胞原代培养及纯化方法的优化探索  

Research for a Better Method of Neonatal Rat Cardiac Myocytes,Primary Culture and Purification

作　　者：蒋学友[1] 张丽[1] 俞晨[1] 江浩[1] 李娟[1] 

机构地区：[1]东南大学第二附属医院急诊科,南京210003

出　　处：《四川大学学报（医学版）》2015年第2期301-304,共4页Journal of Sichuan University(Medical Sciences)

基　　金：南京市医学科技发展资金(No.QYK11158)资助

摘　　要：目的通过对乳鼠心肌细胞原代培养及纯化方法的优化探索,寻找简单、有效的培养及纯化方法。方法 20只乳鼠随机分两组,A组采用酶消化法+吸管吹打获取心肌细胞;B组通过酶消化法+自制转子磁力搅拌获取心肌细胞。两组均采用差速贴壁和5-溴脱氧尿核苷(BrdU)纯化心肌细胞,免疫荧光法鉴定心肌细胞纯度,台盼蓝染色观察心肌细胞活性,显微镜下计数心肌细胞搏动性。结果 A组心肌细胞的存活率、搏动率、纯度分别为(89.90±2.92)%、(91.30±2.00)%和(94.90±1.79)%;B组分别为(94.70±2.31)%、(95.00±1.24)%和(95.60±1.43)%。B组心肌细胞活性及搏动率均高于A组,差异有统计学意义(P>0.05);B组的心肌细胞纯度稍高于A组,但差异无统计学意义(P>0.05)。BrdU加入后与未加入相比,抑制成纤维细胞的生长。结论酶消化法+自制转子磁力搅拌更优于酶消化法+吸管吹打,可以有效获取心肌细胞,BrdU有利于心肌细胞的纯化。Objective To seek a simple and effective approach through probing method about primary culture and purification of neonatal rat cardiac myocytes.Methods Twenty neonatal rats were randomly divided into 2groups.Group A：the cardiac myocytes were gained by means of enzymic digestion and tubularis-blowing;Group B：the cardiac myocytes were gained by means of enzymic digestion and rotator-stirring.The cardiac myocytes in both groups were purified by means of differential attachment technique and BrdU inhibition and were identified with immunofluorescence staining.Viability was assessed with trypan blue staining and pulsation was assessed under microscope.Results The rates of cardiac myocyte＇s viability,pulsation and purity in group A were（89.90±2.92）%,（91.30±2.00）% and（94.90±1.79）%respectively;The corresponding rates in group B were（94.70±2.31）%,（95.00±1.24）% and（95.60±1.43）% respectively.The rates of cardiac myocyte＇s viability,pulsation in group B were significantly higher than that of group A,which was statistically different（P〉0.05）.The rate of cardiac myocytes purity in group B was a bit higher than group A,but which wasn＇t statistically different（P〈0.05）.Fibroblasts cultured with BrdU grew weaker and less than those without BrdU.Conclusion The method of rotator-stirring and enzymic digestion was better than that of tubularis-blowing and enzymic digestion,which helped to gain cardiac myocyte effectively.BrdU helped to purify cardiac myocytes.

关 键 词：乳鼠 心肌细胞 原代培养 

分 类 号：R725.4[医药卫生—儿科]