小鼠大脑皮质星形胶质细胞的原代培养及鉴定  

Primary Culture and Identification of Mouse Cerebral Cortex Astrocytes

作　　者：高帆 余资江[1] 曹文鹏[1] 李玉美[1] Gao Fan;Yu Zijiang;Cao Wenpeng;Li Yumei(Department of Human Anatomy, Guizhou Medical University, Guiyang, 55000)

机构地区：[1]贵州医科大学人体解剖学教研室

出　　处：《基因组学与应用生物学》2018年第7期3149-3152,共4页Genomics and Applied Biology

基　　金：国家自然基金(81060108);贵州省科技厅社会发展攻关(黔科合SY字(2012)3153号)共同资助

摘　　要：为了体外分离培养小鼠大脑皮质星形胶质细胞（astrocyte,AS）。取新生24 h内ICR乳鼠,超净台内断头取脑,体视显微镜下剥除脑膜、血管及海马,获得完整的大脑皮层组织,经手术刀切碎组织,旋涡震荡,过两次尼龙筛网以制备单细胞悬液,接种于35 mm塑料培养皿,倒置相差显微镜下观察细胞形态学状况,培养至3~4周,进行星形胶质细胞标记性蛋白胶质纤维酸性蛋白（glial fibrillary acidic protein,GFAP）免疫荧光鉴定细胞纯度。本研究表明,倒置相差显微镜下观察细胞于接种后2~3 d大部分贴壁,生长至7 d左右即铺满皿底,3~4周细胞生长成熟,胞体较大,形状不规则,呈“铺路石”（cobblestone-like structure）状,免疫荧光鉴定可见GFAP阳性细胞占细胞总数比例达95%以上。通过以上方法可获得高纯度星形胶质细胞,为下一步实验提供大量生长状态良好的细胞。To isolate and culture the astrocytes from mouse cortex in vitro. The newbom ICR mice within 24 h were used to the bilateral cortex in clean bench. The meninges, blood vessels and the hippocampus were removed from cortices under stereomicroscope. The cortices were minced with surgical blade, shocked with oscillator, filtered with nylon mesh twice to obtain the single-cell suspension. Then the cells were inoculated in 35 mm plastic dishes, and the cells were observed under inverted phase contrast microscope. 3-4 weeks after being cultured, purity of the cells were identified with immunostaining of astrocyte marker, GFAP. This research showed that after inoculated 2-3 d the cells sticked wall most, grown to about 7 d, the cells were paved with the bottom of dishes, 3-4 week, the cells grown mature, shape irregularity, cobblestone-like structure, immunofluorescence identification of GFAP positive cell rate of more than 95% of total ceils. Through the above method can gain high purity astrocytes, for the next experiment provides a number of growth of cells in good status.

关 键 词：星形胶质细胞 原代培养 GFAP 

分 类 号：R338[医药卫生—人体生理学]