一种大鼠腹腔肥大细胞分离方法  

A method for isolating rat peritoneal mast cell

作　　者：幸晓燕[1] 王青[1] 周联[1] 邓向亮[1] 

机构地区：[1]广州中医药大学临床药理研究所免疫研究室,510405

出　　处：《免疫学杂志》2011年第7期615-618,624,共5页Immunological Journal

基　　金：广东高校优秀青年创新人才培育项目(LYM09049);广州省中医药管理局科研项目(20092251)

摘　　要：目的介绍一种密度梯度离心分离大鼠腹腔肥大细胞(RPMC)的技术。方法比较改良方法和原方法2种分离的细胞量、活性及纯度。台盼蓝染色检测细胞存活率;甲苯胺蓝染色检测肥大细胞纯度。将弃去密度梯度离心上液2 m(lA方法)与2.5 m(lB方法)比较,以分析弃去的细胞液的量与肥大细胞纯度的关系。用C48/80刺激分离的RPMC,检测细胞释放的β-氨基己糖苷酶和组胺以确定分离的RPMC脱颗粒活性;透视电镜扫描观察细胞。结果改良方法分离的细胞显著多于原方法分离的细胞数;两种方法分离的细胞存活率都在99%以上;肥大细胞纯度都在90%以上。C48/80刺激RPMC,分离的细胞释放大量的β-氨基己糖苷酶和组胺,高于空白对照组。电镜观察细胞具有明显的肥大细胞形态。结论改良的Percoll密度梯度离心分离大鼠腹腔肥大细胞的方法可行。Lennart had introduced a method of isolation rat peritoneal mast cell（RPMC） in 1980,which,however,do not widely used today.Here,we introduce an optimized method,which was compared with old method（Lennart method） in this study.Trypan blue indicator solution and toluidine blue indicator solution indicated that the viability and purity of mast cell isolated by the two methods similar.The relation of the discarded suspension quantity with the purity of mast cell in the two methods was analyzed with the same methods used before,and we found that 2.5 ml discarded suspension could make high purity of RPMC in both methods.Compound 48/80 was used to stimulate RPMC,and then the degranulation was detected by measuring released hexosaminidase and histamine quantity.Furthermore,electron microscope demonstrated that the cells are mast cell appearance.All these come to that the optimized method is feasible.

关 键 词：腹腔肥大细胞 PERCOLL C48/80β-氨基己糖苷酶 组胺 甲苯胺蓝 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]