SD大鼠的胎鼠海马神经元细胞的分离培养和鉴定  

Isolation,cultivation and identification of hippocampal neurons from prenatal mice

作　　者：张文雅[1] 程秀永[1] 张静[1] 王丹丹[1] 孙素珂 

机构地区：[1]郑州大学第一附属医院新生儿科,450052 [2]河南省高等学校临床医学重点学科开放实验室,郑州450052

出　　处：《中国实用医刊》2017年第10期1-4,F0004,共5页Chinese Journal of Practical Medicine

基　　金：国家自然科学基金（81471491）

摘　　要：目的 建立一种理想的体外海马神经元原代培养的实验方法.方法 取孕18 d的SD大鼠,分离体内胎鼠的海马后,经木瓜蛋白酶消化后接种,实验前用多聚赖氨酸铺板、采用无血清培养海马神经元,并采用免疫荧光进行神经元特异性标志物的鉴定;利用MTT 比色法分析检测神经元细胞在不同处理方式下的成活率.结果 海马神经元生长状态良好,培养24 h后可形成突触,7-12 d可形成成熟的神经细胞网络,经NSE免疫荧光鉴定神经元纯度达95%以上.消化时采用木瓜蛋白酶比胰蛋白酶的细胞成活率明显增高（P〈0.01）;采用口径1 mm枪头吹打细胞比口径0.5 mm和200目筛的细胞成活率高（P〈0.01）.结论 该实验方式可获得高纯度的海马神经元细胞,并为一种简单易行的细胞培养方法.Objective To establish a favorable primary culture of hippocampal neurons isolated from prenatal mice.Methods The hippocampus of embryonic SD mouse aged 18 d was isolated,digested with papain,and then the neurons were cultured in planks with polylysine before experiment,we used Neurobasal as a culture medium for neurons which was without serum.And the specific markers of neurons were identified through immunofluorescence using MTT colorimetric analysis to detect the survival rate of neuronal cells under different treatments.Results Hippocampal neurons grow well,synapses grow out after 24 hours from cultivation and a mature network of nerve cells can be formed after 7-12 d.The NSE immunofluorescence identified neurons of more than 95% purity.Compared with trypsin,the cell survival rate was higher with papain in digestion （P〈0.01）;The cell survival rate of normal tip-1.0 mm was higher than tip-0.5 mm and 200 mesh sieve （P〈0.01）.Conclusions By this technique,we can obtain hippocampal neurons with high purity,and it is a simple,effective method for cell culture.

关 键 词：海马神经元 原代培养 SD胎鼠 

分 类 号：Q813.1[生物学—生物工程] R-33[医药卫生]