一种乳鼠海马神经元原代培养方法的优化与鉴定  

Optimization and Identification of Primary Culture Method of Neonatal Rat Primary Hippocampal Neurons

作　　者：吴倩[1,2,3] 穆婷婷 汪宁[1,2,3] Qian Wu;Tingting Mu;Ning Wang(Anhui University of Chinese Medicine,Anhui Hefei 230012,China;Anhui Province Key Laboratory of Research&Development of Chinese Medicine,Anhui Hefei 230012,China;Anhui Academy of Traditional Chinese Medicine-Institute for Pharmacodynamics and Safety Evaluation of Chinese Medicine,Anhui University of Chinese Medicine,Anhui Hefei 230012,China)

机构地区：[1]安徽中医药大学,安徽合肥230012 [2]中药复方安徽省重点实验室,安徽中医药大学,安徽合肥230012 [3]安徽省中医药科学院中药药效与安全评价研究所,安徽中医药大学,安徽合肥230012

出　　处：《中医药临床杂志》2021年第5期923-926,共4页Clinical Journal of Traditional Chinese Medicine

基　　金：国家自然科学基金青年项目“基于Parkin介导的线粒体自噬研究藁本内酯治疗缺血性脑损伤的机制”(项目编号:81903954);安徽高校自然科学研究重点项目“基于AMPK对海马神经元线粒体质量控制的靶向调节探讨藁本内酯防治缺血性脑卒中的作用机制”(项目编号:KJ2020A0384)。

摘　　要：目的:建立一种稳定高效、纯度较高的大鼠海马神经元体外原代培养的方法,并用免疫荧光法对其进行纯度鉴定,为缺血性脑卒中研究的体外模型建立提供一定的实验技术参考。方法:取新生24h的SD大鼠,经消毒、断头取脑分离出海马组织,剪碎后经过消化、重悬,并用含10%血清的DMEM/F12培养基进行接种培养,于第2天半量换为Neurobasal+B27培养基,后每2天换液1次。倒置显微镜下观察不同时期神海马经元生长过程的形态学变化。采用MAP-2、Cy3神经元特异性抗体对其进行免疫荧光鉴定。结果:体外培养海马神经元结构特征明显,生长状况良好,能形成典型的神经细胞网络,细胞鉴定呈阳性,纯度较高。结论:上述改良后海马神经元的原代培养,可获得生长状态良好的海马神经元,是体外研究缺血性脑卒中的良好细胞模型。Objective:To establish a stable,efficient and high purity primary culture method of rat hippocampal neurons in vitro,and to identify their purity by immunofluorescence,so as to provide a certain experimental technical reference for the establishment of in vitro model in the study of ischemic stroke.Methods:Newborn SD rats were taken for 24h,disinfected and decapitated,and the hippocampal tissues were isolated.After being cut into pieces,the hippocampal tissues were digested and resuspended.DMEM/F12 medium containing 10%serum was used for inoculation and culture.The morphological changes of the growth process of the hippocampus at different stages were observed under an inverted microscope.They were identified by immunofluorescence with MAP-2 and Cy3 neuron-specific antibodies.Results:Hippocampal neurons cultured in vitro had obvious structure characteristics,good growth status,and could form typical neural cell network.Conclusion:The improved primary culture of hippocampal neurons can obtain hippocampal neurons with good growth state,which is a good cell model for studying ischemic stroke in vitro.

关 键 词：海马神经元 原代培养 优化和鉴定 

分 类 号：R-331[医药卫生]