脂质体转染人肝星状细胞和肝细胞条件优化

Optimization of the Transfection Efficiency of Human Hepatic Stellate Cell and Hepatocyte Using Liposome

利用Lipofectamine 2000（Lipo）介导LacZ及EGFP基因转染人肝星状细胞系LX-2和人肝细胞系Chang Liver,探讨Lipo用量、质粒用量、转染时间、细胞初始接种密度及不同启动子驱动对转染效率的影响.结果显示,两种细胞均能得到高效转染,Chang Liver细胞转染效率（＆gt;80%）高于LX-2细胞（＆gt;60%）.Lipo用量为每孔2μL,转染后6 h换液,LX-2细胞应用质粒DNA为每孔0.45μg,Chang Liver细胞应用质粒DNA为每孔0.30~0.45μg,此时转染效率最高.巨细胞病毒（CMV）启动子驱动LacZ转染效率与CMV早期增强子/鸡β肌动蛋白（CAG）启动子驱动增强型绿色荧光蛋白（EGFP）转染效率无显著差异;细胞初始接种密度对转染效率无显著影响.

PAAV- CMV-LacZ plasmid and pAAV-CAG-EGFP plasmid were transfected into a kind of human hepatic stellate cell line-LX-2 and a kind of human hepatocyte line-Chang Liver using Lipofectamine 2000, respeetivelyo The impacts of liposome and plasmid amount, transfection time course, initial plated density and different promoter used on transfeetion efficiency were discussed. It indicated that both kinds of cell lines were highly transfeetable, and the transfec- tion efficiency of Chang Liver cells （〉80%） was higher than LX-2 cells （〉60%）. The transfection efficiency turned out to be the highest while using 2 μL· well-1 of liposome, 0. 45 μL· well-1 of plasmid DNA for LX-2 cells and 0. 30-0. 45μL·well-] of plasmid DNA for Chang Liver cells after 6 hours of transfection. There was no significantly difference in transfection efficiency whether the gene was driven by CMV or CAG promoter. The initial plated density seemed to have no obvious influence on the transfection efficiency, either.