两种人卵巢颗粒细胞分离提纯方法的比较研究  

Comparison of Two Methods for Isolation and Purification of Human Granulosa cells

作　　者：江欢[1] 谢宝国[1] 罗琛[2] 李红[2] 朱伟杰[1] 

机构地区：[1]暨南大学生命科学技术学院,生殖免疫研究所,广州510632 [2]南方医科大学南方医院妇产科生殖医学中心,广州510515

出　　处：《生殖与避孕》2012年第11期728-732,共5页Reproduction and Contraception

摘　　要：目的:比较2种人卵巢颗粒细胞分离提纯方法的效果。方法:收集体外受精-胚胎移植(IVF-ET)穿刺取卵后的卵泡液,分别用2种方法分离提取颗粒细胞。A法用红细胞裂解液去除混杂红细胞、酶消化卵泡液中黏液团、Percoll分离液与细胞悬液等体积叠加后离心;B法省略红细胞裂解步骤、舍弃黏液团、增加Percoll分离液用量。比较2种方法获取的颗粒细胞数量、活性、纯度以及实验耗时。结果:2种方法获取的颗粒细胞存活率均>90%,差异无统计学意义(P>0.05)。A法获取的颗粒细胞数量多于B法(P<0.05),但混杂较多细胞碎片,体外培养细胞生长状态欠佳;B法获取的颗粒细胞纯度高,体外培养生长状态良好,且实验耗时较A法短(P<0.05)。结论:2种方法对颗粒细胞存活率无明显影响。B法更为省时高效,有助于建立稳定的颗粒细胞体外培养体系。Objective： To compare two methods for isolation and purification of human granulosa cells. Methods： Follicular fluid was collected after ovary extraction through puncture in IVF-ET and was used to seperate human granulosa cells by 2 different methods which divided into group A and group B. In group A, samples were prepared by erythrocyte lysis, digestion of cumulus cells and centrifugation with isometric Percoll. In group B, the method was simplified in terms of omission of erythrocyte lysis, discard of mucus cumulus and increased doses of Percoll. These two methods were fully validated for yield, purity and viability of granulosa cells and time consuming. Results： Viability of granulosa cells was similar between group A and group B （P〉0.05）. Group A was characterized by its better yield of granulosa cells （P〈0.05）, yet accompanied with increased cell debris and cell lose in long-term culture. The method used in group B led to high in vitro viability and purity of granulosa cells, and in addition allowed a more time-consuming procedure （P〈0.05）. Conclusion： These two methods have no impact on viability of the isolated granulosa cells, whereas the simplified method applied in group B is more efficient and time-saving and this is of great value for the establishment of a stable in vitro culture system of human granulosa cells.

关 键 词：卵巢颗粒细胞 细胞分离 体外培养 

分 类 号：R321.1[医药卫生—人体解剖和组织胚胎学]