BALB/c鼠肝脏枯否细胞不同分离培养方法的比较  

The isolation and culture of kupffer cell with different methods in BALB/c mouse

作　　者：柳俊刚[1] 高枫[2] 张森[2] 杨剑锋[1] 刘传渊[1] 

机构地区：[1]广西医科大学研究生学院,南宁530021 [2]广西医科大学第一附属医院结直肠外科,南宁530021

出　　处：《结直肠肛门外科》2009年第5期311-314,共4页Journal of Colorectal & Anal Surgery

基　　金：国家自然基金项目编号30560150;30672059;广西医疗卫生重点科研课题项目编号重200404

摘　　要：目的建立简便易行,存活好、产量和纯度高,稳定的枯否细胞(Kupffer Cell,KC)分离培养方法。方法根据酶消化时间不同和裂解红细胞的方法在酶消化之后和之前分方法Ⅰ和方法Ⅱ。分别运用0.1%的Ⅱ型胶原酶、0.05%链酶蛋白酶E和0.005%DnaseⅠ在不同方法中离体消化肝组织并进行密度梯度离心和贴壁培养提取KC,通过吞噬实验观察鉴定KC。结果方法Ⅰ和方法Ⅱ中KC的数量分别是(0.71±0.21)×107/g和(0.96±0.20)×107/g,细胞贴壁率分别40.1%、45.1%。用0.4%台盼蓝染色鉴定,存活率分别为90%、95%。吞噬实验发现,细胞纯度分别为90%、95%。结论结合碾磨筛网滤过降低酶作用时间并在酶消化之前裂解红细胞分离提取枯否细胞具有存活好产量纯度高的优点。Objective To establish a simple,activity and high-yield isolation method of Kupffer cells（KC）.Methods Two methods（ⅠandⅡ） were used by the times of enzymatic digestion and cracking red blood cells after or before.KC were isolated by 0.1%CollagenaseⅡ,0.05% Pronase E and 0.005% DnaseⅠdigestion of liver,density gradient centrifugation and adherent culture,identified by the phagocytosis of ink.Results The number of KC by methodsⅠand Ⅱ were（0.71±0.21） ×10^7/g and（0.96± 0.20） ×10^7/g,cell adhesion rates were 40.1% and 45.1%. Identification by 0. 4% trypan blue shows the cells survival rates were 90% and 95%. The cells purity were 90% and 95% in phagocytosis. Conclusion Combination of milling-screen filter to reduce the digestive time of enzyme and crack RBC before enzymes digestion is a good method for isolating KC.

关 键 词：枯否细胞 分离 鉴定 酶消化法 

分 类 号：R575[医药卫生—消化系统]