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目的 建立HBV感染人胎肝细胞体外培养系统。 方法 首先分离、培养人胎肝细胞,然后应用HBV阳性血清感染体外培养的人胎肝细胞;每隔2d收集上清液和肝细胞,应用ELISA、免疫组化法、原位杂交法和斑点杂交法检测上清液和细胞中HBsAg和HBV DNA。 结果 上清液中HBsAg在感染后2d～20d均可测出,以感染后4d～16d达高峰(A值在0.22左右)。免疫组化检测细胞中HBsAg呈阳性表达,原位杂交和斑点杂交检测细胞和上清液中HBV DNA也呈阳性表达。 结论 HBV在原代培养人胎肝细胞中能稳定复制和表达至少达16d。
Objective To establish a HBV-infected human fetal hepatocyte culture system in vitro. Methods Human fetal liver cells were isolated and cultured. HBV-positive human sera were used to infect human fetal hepatocytes cultured in vitro. Supernatants and hepatocytes were collected every 2 days. ELISA, immunohistochemistry, in situ hybridization and dot blot Detection of HBsAg and HBV DNA in supernatants and cells. Results HBsAg in the supernatant was detected at 2 days to 20 days after infection and peaked at 4 days to 16 days after infection (A value was about 0.22). Immunohistochemical detection of HBsAg positive cells, in situ hybridization and dot blot hybridization detection of HBV DNA in the cells and supernatant was also positive. Conclusion HBV can be stably replicated and expressed in primary cultured human fetal hepatocytes for at least 16 days.