大鼠肝陷窝细胞分离、鉴定及纯度、活性分析  

Isolation,Purification and Identification of Pit Cells in Rat Liver

作　　者：和春肖[1,2] 谢来峰[1,2] 王磊[1,2] 樊晋宇[2] 徐存拴 

机构地区：[1]河南师范大学生命科学学院,河南新乡453007 [2]河南省-科技部共建细胞分化调控重点实验室,河南新乡453007

出　　处：《河南科学》2009年第9期1072-1076,共5页Henan Science

基　　金：河南省重大公益性科研计划项目(081100910700)

摘　　要：按Higgins等方法制作大鼠2/3肝切除(parital hepatectomy,PH)模型,用两步灌流法分散肝脏细胞,用60% Percoll梯度离心结合免疫磁珠方法分离陷窝细胞(pit cell),用分化抗原簇8(cluster of differentiation8,CD8)和分化抗原簇56(cluster of differentiation 56,CD56)的免疫组织化学方法定性、定位再生肝(regenerating liver,RL)、分散的肝脏细胞及分离的肝陷窝细胞,用RT-PCR定量肝陷窝细胞的CD8和CD56 mRNA,用蛋白免疫印迹定量肝陷窝细胞的CD8和CD56蛋白.初步证实,分离的肝陷窝细胞中,CD8和CD56阳性细胞占95%以上;PH后各时间点分离的陷窝细胞的CD8和CD56的mRNA量稳定,相应的蛋白量亦稳定.表明改进的分离陷窝细胞方法具有收率和纯度高、活性好等特点,值得采用.Rat 2/3 hepatectomy （PH） model was done following Higgins et al, hepatic cells were scattered by twostep perfusion, and pit cells were isolated and purified by density gradient centrifugation with 60 % percoll and immunomagnetic beads. Immunocytochemistry method was used to qualitify and localize cluster of differentiation 8 （CD8） and cluster of differentiation 56 （CD56） in liver tissue, the isolated hepatic cells, and the purified pit ceils. The exrpressions of CD8 and CD56 were quantified using RT-PCR. The results showed that CD8 and CD56 postive pit ceils account up more than 95 % of the total pit cells; mRNA levels of CD8 and CD56 in the isolated pit cells were stable in rat regenerating liver （RL）, and also was the content of the corresponding proteins, indicating the modified method for pit cells isolation in this study has the advantage of high pit cell harvest, high purification and survival rate.

关 键 词：大鼠 细胞分离和鉴定 陷窝细胞 免疫磁珠 标记蛋白 

分 类 号：Q2-33[生物学—细胞生物学]