摘要
目的:观察针对HBV X的siRNA表达载体pGenesil-HBV X与阳离子脂质体的复合物对HepG2.2.15细胞转染效果及其对HBVM表达的影响。方法:以pGenesil-siAFP表达载体和非转染细胞分别作非特异性序列对照组和空白对照组,用不同配比浓度pGenesil-HBV X与阳离子脂质体的复合物转染HepG2.2.15细胞,荧光显微镜下观察并计算转染阳性细胞百分率,筛选出最佳转染效率的剂量配比。于转染后24h、48h、72h采用时间分辨荧光免疫测定技术(TRFIA)检测上清液中HBsAg和HBeAg。结果:pGenesil-HBV X与阳离子脂质体复合物8μL∶2.5μg剂量配比组的平均转染率达到了(55.1±4.3)%,且不影响细胞活性;转染后48h和72h后该siRNA表达载体对HepG2.2.15细胞表达HBsAg和HBeAg抑制作用显著(P<0.05)。结论:8μL:2.5μg剂量配比组的pGenesil-HBV X与阳离子脂质体复合物是最佳的转染剂量配比,能有效抑制HepG2.2.15细胞表达HBsAg和HBeAg。
Objective： To observe the ratios of pGenesil - HBV X and lipid - complex （Metafectene） transfecting HepG2.2.15 cells, and the effect of inhibiting HBVM expression. Methods： Control groups： pGenesilsiAFP and non - transinfected HepG2.2.15 cells. Transinfecting HepG2.2.15 cells by using different ratios of pGenesil - siAFP and Metafectene, to caculate the percents of trasinfected cells by fluoroscope and get the proper ratio of the angent. After 24h, 48h, 72h, the expression of HBV surface antigen （HBsAg） and e antigen （HBeAg） secreting into the supernatant were determined with Time - resolved Immunofluorometric Assay kit （TRFIA） . Results ： The proper ratio of pGenesil - siAFP and Metafectene is 8μL ： 2.5μg （55.1 ± 4.3） % cells transinfected）, pGenesil - HBV X could significantly reduce HBV antigen expression in supernatant of HepG2. 2.15 after transfection 48h and 72 h（ P 〈 0.05）. Conclusions ： 8μL ： 2.5μg is proper ratio of pGenesil - siAFP and Metafectene transinfecting cells, which inhibited HepG2.2 15 expressing HBVM
出处
《长治医学院学报》
2005年第3期161-164,共4页
Journal of Changzhi Medical College
基金
山西省自然基金研究项目(20051114)
