壁虎胚胎大脑皮层神经元和胶质细胞的分离、培养及纯化

目的 建立多疣壁虎胚胎大脑皮层神经元和胶质细胞的分离、培养及纯化方法.方法 分离孵化15d的多疣壁虎胚胎的大脑皮层,经胰酶消化,细胞计数后接种于培养瓶.采用差速贴壁及反复传代相结合的纯化培养方法培养胶质细胞;采用添加B27的神经元培养基培养神经元,并用免疫细胞化学方法鉴定.结果 获得体外培养的壁虎GFAP阳性表达胶质细胞,4代后纯度达95%以上;采用神经元培养基,神经元生长良好,NF、MAPⅡ表达阳性,第10d纯度达95%以上.结论 建立了壁虎细胞的培养方法,并获得高纯度的胶质细胞和神经元,为进一步对神经系统的深入研究提供细胞模型.

ISOLATION, CULTURE AND PURIFICATION OF CEREBRAL CORTICAL NEURONS AND GLIAL CELLS FROM THE EMBRYONIC GEKKO

Objective To establish a method of neurons and glial cell culture from embryonic Gekko japonicus cerebral cortex. Methods Embryonic(E15) pallium was dissociated and digesting by trypsin.After counting,cells were seeded in culture flask.The glial cells were obtained by using differential adhesion potential combined with successive passage purified methods,and neurons were obtained by using neurobasal medium supplemented with B27.The cells were fixed and analyzed with immunohistochemic assay. Results After tetra-generation,GFAP positive cells were more than 95% in glial cells cultured condition;Neurons grew well in neurobasal medium,and NF and MAP-2 positive signals co-localized on neurons.After cultured for 10 days,the percentage of neurons was more than 95%.Conclusion The methods of isolation,culture and purification for embryonic Gekko japonicus cortical neurons and glial cells were established and it might be a valuable cell model to further investigate the central nerve system in Gekko japonicus.