Wang An-hui, Men Ke, Yan Yong-ping, Xu De-zhong, Lu Juan, Wang Xue-ping, Zhang Jing-xia
Department of Epidemiology, Fourth Military Medical University, Xi'an 710032, China.
Zhonghua Fu Chan Ke Za Zhi. 2006 Mar;41(3):165-8.
To determine the role of hepatitis B Immunoglobulins (HBIG) in blocking hepatitis B virus (HBV) infection of trophoblast cell culture in vitro.
Trophoblast cells were placed in the six-well cluster dishes and incubated with 10% fetal calf serum/Dubecco's modified Eagle's Medium (10% FCS DMEM) at 37 degrees C with 5% CO2 in air. At 24 h after plating cells were subjected to experiment. Group A: cells were cultured with 0.5 ml HBV positive serum plus 3 ml 2% FCS DMEM; Group B: cells were cultured with 3 ml 2% FCS DMEM plus 0.5 ml HBV positive serum pretreated with 80 U HBIG for 30 min at 37 degrees C; Group C: cells were cultured with 3 ml 2% FCS DMEM plus 0.5 ml HBV positive serum pretreated with 40 U HBIG for 30 min at 37 degrees C; Group D: cells were cultured with 3 ml 2% FCS DMEM plus 40 U HBIG for 30 min before 0.5 ml HBV positive serum was added; Group E: cells were cultured with 40 U HBIG plus 3 ml 2% FCS DMEM; Group F: cells were cultured with HBV negative serum plus 3 ml 2% FCS DMEM. Twenty-four hours later the inoculums were removed, and the cells were extensively washed with 0.01 mol/L phosphate-buffered saline (PBS). After PBS washing, 4 ml 2% FCS DMEM was added to each well and the medium was collected every 12 hours. Enzyme-Linked Immunosorbent Assay (ELISA) method was used to detect HBsAg in culture medium (absorption value, A). HBV DNA in cell culture medium was detected by polymerase chain reaction (PCR).
Before PBS washing, the A value of groups A, B, C, D, E, F were 2.697, 0.040, 0.102, 0.198, 0.036, 0.040 respectively. The cell culture medium in groups of A, B, C, and D were HBV DNA positive, groups of E, F were HBV DNA negative. From 12 hours to 84 hours, the average A value of groups A, B, C, D, E and F was 1.55 +/- 0.27, 0.032 +/- 0.016, 0.100 +/- 0.087, 0.052 +/- 0.044, 0.034 +/- 0.020, 0.034 +/- 0.022 respectively. The A value of groups A was significantly higher than those of other groups (P < 0.01). Cell culture medium at 84 hours of group A was HBV DNA positive and those of group B, C, D, E, F were HBV DNA negative.
HBIG could effectively block HBV infection of trophoblast cell culture in vitro.
确定乙肝免疫球蛋白（HBIG）在体外阻断滋养层细胞培养中乙肝病毒（HBV）感染的作用。
将滋养层细胞置于六孔板中，在含5%二氧化碳的空气中，于37℃用10%胎牛血清/杜氏改良伊格尔培养基（10% FCS DMEM）培养。接种细胞24小时后进行实验。A组：细胞用0.5 ml HBV阳性血清加3 ml 2% FCS DMEM培养；B组：细胞用3 ml 2% FCS DMEM加0.5 ml在37℃用80 U HBIG预处理30分钟的HBV阳性血清培养；C组：细胞用3 ml 2% FCS DMEM加0.5 ml在37℃用40 U HBIG预处理30分钟的HBV阳性血清培养；D组：细胞在加入0.5 ml HBV阳性血清前先用3 ml 2% FCS DMEM加40 U HBIG培养30分钟；E组：细胞用40 U HBIG加3 ml 2% FCS DMEM培养；F组：细胞用HBV阴性血清加3 ml 2% FCS DMEM培养。24小时后吸出接种物，用0.01 mol/L磷酸盐缓冲盐水（PBS）充分洗涤细胞。PBS洗涤后，每孔加入4 ml 2% FCS DMEM，每12小时收集培养基。采用酶联免疫吸附测定（ELISA）法检测培养基中的HBsAg（吸光度值，A）。用聚合酶链反应（PCR）检测细胞培养基中的HBV DNA。
PBS洗涤前，A、B、C、D、E、F组的A值分别为2.697、0.040、0.102、0.198、0.036、0.040。A、B、C、D组的细胞培养基HBV DNA阳性，E、F组的HBV DNA阴性。从12小时至84小时，A、B、C、D、E、F组的平均A值分别为1.55±0.27、0.032±0.016、0.100±0.087、0.052±0.044、0.034±0.020、0.034±0.022。A组的A值显著高于其他组（P<0.01）。84小时时A组细胞培养基HBV DNA阳性，B、C、D、E、F组的HBV DNA阴性。
HBIG可有效阻断体外滋养层细胞培养中的HBV感染。
