一种高效简便的原代神经元培养方法  

An Efficient and Convenient Model of Culturing Primary Neurons

作　　者：张涛[1,2] 胡怀强[1] 王旭辉[3] 牛兵[1] 陶珍[1] 曹秉振[1] ZHANG Tao HU Huai-qiang WANG Xu-hui NIU Bing TAO Zhen CAO Bing-zhen(General Hospital of Jinan Military Area, Jinan 250031, China No.303 Hospital of Chinese People＇s Liberation Army, Nanning 530021, China Daping Hospital, Research Institute of Surgery Third Military Medical University, Chongqing 400042, China)

机构地区：[1]济南军区总医院,济南250031 [2]解放军第303医院,南宁530021 [3]第三军医大学大坪医院野战外科研究所,重庆400042

出　　处：《神经损伤与功能重建》2016年第6期471-472,475,共3页Neural Injury and Functional Reconstruction

基　　金：十二五全军重点项目No.BWS11J062

摘　　要：目的：建立一种简便高效的原代神经元培养方法。方法：出生12 h内的Wistar乳鼠,分离皮质后剪碎,0.25%胰酶消化30 min,漂洗后轻柔吹散细胞,过滤后1 000 rpm离心5 min,弃上清,加接种液吹匀后过滤,静置3～5 min,取上层液体计数并接种。接种后4、45、96 h全量换液,之后每3 d半量换液1次。培养第5天评估神经元,NSE染色、NMDAR1染色和台盼兰染色。结果：神经元纯度高（〉95%）,死亡率低（〈5%）,细胞形态好,交联充分,背景干净。结论：利用新生大鼠皮质建立了高质量、高产量、简便的神经元培养模型。Objective： To establish an efficient and convenient model of culturing primary neurons. Methods：Firstly, the cortex of newborn Wistar mice younger than 12 hours old was thoroughly separated. And then the cortex was digested by 0.25% tyrisin for 30 min after cutting into pieces. Cells were rinsed, blew and centrifuged at1000 rpm for 5 min after filtering. The supernatant was abandoned, and the resultant was added with plantation medium, gently blew and filtered. After standing for 3～5 min, the supernatant liquid was withdrawn in order to perform a count on the cells. The medium was replaced in full at 4 h, 45 h and 96 h later after seeding and then the medium could be replaced in half every 3 days. At day 5 of culture, the neurons were evaluated by NSE, NMDAR1 and trypan blue staining. Results： The purity of neurons was over 95% and the mortality rate was less than 5%. The morphology of neurons was excellent, effectively crosslinking, and the background was clean. Conclusion： A high-quality, high-yield and simple neuronal culture model was established using neonatal rat cortex.

关 键 词：皮质神经元 原代培养 细胞模型 乳鼠 

分 类 号：R741[医药卫生—神经病学与精神病学] R741.02[医药卫生—临床医学]