微小磁珠法分离胚胎大鼠神经干细胞的实验研究  

Primary Study to Separate the Neural Stem Cells from the Fetal Rat Brain by Using the Immunomagnetic Microbeads

作　　者：王彦惠[1] 刘振华[1] 廖可立[2] 姜晓丹[2] 李留洋[3] 徐如祥[2] 邹雨汐[2] 戴宜武[2] 杜谋选[2] 蔡颖谦[2] 

机构地区：[1]第一军医大学珠江医院神经内科,广东广州510282 [2]第一军医大学珠江医院神经外科,广东广州510282 [3]第一军医大学珠江医院肾移植科,广东广州510282

出　　处：《中国神经免疫学和神经病学杂志》2002年第4期196-199,F003,共5页Chinese Journal of Neuroimmunology and Neurology

基　　金：军队"十五"重点项目基金资助项目(0 1ZD5 4);广东省科技厅重点项目基金资助项目[粤科基办 (2 0 0 0 ) 2 5 ;粤财企 (2 0 0 1) 3 67]

摘　　要：目的　旨在以微小免疫磁珠 (平均直径≤ 5 0 nm)分离提纯 SD大鼠胚胎大脑皮层组织中的神经干细胞并进行培养 ,观察细胞的增殖分化情况并对其分化产物进行鉴定 ,评价该分离方法的可行性及其在神经干细胞研究中的应用价值。方法　取 SD大鼠的胚胎大脑皮层组织制单细胞悬液 ,用微小磁珠分选出神经巢蛋白(nestin)阳性的细胞群 ,光镜下检测其纯度及活力后进行体外培养 ,特定条件下诱导分化并以免疫细胞方法检测产物的细胞类型。结果　分离的神经干细胞纯度为 (89.2± 4.8) % ,细胞存活率 (97.0± 1 .6) % ,诱导分化产物中出现 NSE阳性、GFAP阳性、巢蛋白 (nestin)反应阳性细胞群落。结论　微小磁珠法分离提纯神经干细胞纯度高 ,获得细胞数多 ,并对细胞的生物活性无明显影响 。Objective The objective of the study is to separate the neural stem cells from the fetal rat brain by immunomagnetic beads (d≤50 nm) technique then were cultured in vitro, for investigating the possibilities of these cells to differentiate into neuronal cells in vitro.Methods Cell suspension of the fetal rat cerebral cortex was incubated with antibody of nestin and these labeled cells were separated from the suspension through the magnetic field by immunomagnetic beads coated with the second antibody. The purity of the separated cells and the active cells were determined. Then culture these cells in vitro and dissociated them with serum of fetal bovine for 3 days. Immunocytochemistry was performed to identify the properties of the differentiated cells. Results The purity of the separated cells was (89.2±4 8)%, the activity of the cells was accounted for (97 0±1 6)% The treated cells expressed neural proteins as NSEandGFAP. Conclusions The magneticcell sorting system we used can effectively separate neural stem cells from the cerebral cortex cell suspension and maintaining the activity of the cells. The sorted cells are well provided for the subsequent experiments.

关 键 词：微小磁珠法 实验研究 免疫磁珠 神经干细胞 神经巢蛋白 细胞分离 

分 类 号：Q2－33[生物学—细胞生物学]