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目的初步评价肠道病毒71型(Enterovirus 71,EV71)与甲型肝炎病毒(Hepatitis A virus,HAV)灭活疫苗联合免疫小鼠诱导的特异性免疫应答水平以及两种抗原间的相互作用。方法将HAV灭活疫苗和EV71灭活疫苗以不同剂量配比制成联合疫苗,免疫ICR小鼠后,通过ELISA法检测血清HAV抗体效价;固定病毒-稀释血清法检测血清EV71中和抗体效价;流式细胞术检测脾脏中淋巴细胞亚群CD4+/CD8+的百分率;ELISPOT法检测脾脏中淋巴细胞分泌IL-4和IFNγ的水平。结果初免后28 d和加强免疫后7 d,各组小鼠血清中EV71中和抗体阳转率均达100%,单价疫苗组和联合疫苗组EV71中和抗体效价均呈剂量依赖性;初免后28 d,小鼠血清中能够产生保护性HAV抗体,加强免疫后抗体水平升高;联合疫苗组与相应的单价疫苗组间抗体水平差异无统计学意义(P>0.05),未发现EV71与HAV的相互作用;初免后28 d,各组小鼠脾脏中CD8+细胞比例高于CD4+细胞(P<0.001),加强免疫后7 d则相反(P<0.001);初免后28 d,HAV全病毒和EV71多肽均不能刺激各组小鼠脾细胞产生阳性反应,加强免疫后7 d,各组小鼠脾细胞分泌IL-4的水平均高于IFNγ(P<0.05)。结论一定剂量的HAV和EV71联合疫苗可诱导小鼠产生良好的体液免疫应答,同时诱导细胞免疫应答,但相对较弱,未发现两种抗原间存在相互干扰或协同作用。
Objective To evaluate the level of specific immune responses induced by combined immunization of Enterovirus 71 (EV71) and Hepatitis A virus (HAV) inactivated vaccines and the interaction between the two antigens. Methods HAV inactivated vaccine and EV71 inactivated vaccine were used in combination with different dosages to make a combined vaccine. After immunization with ICR mice, the titer of serum HAV antibody was detected by ELISA. The neutralizing antibody of serum EV71 was tested by fixed virus-diluted serum The percentage of CD4 + / CD8 + of lymphocyte subsets in spleen was detected by flow cytometry. The level of IL-4 and IFNγ secreted by lymphocytes in spleen was detected by ELISPOT. Results The positive rate of EV71 neutralizing antibody in serum of mice in each group was up to 100% on the 28th day after vaccination and on the 7th day after booster immunization. The titer of EV71 neutralizing antibody in both monovalent vaccine group and combination vaccine group was dose - dependent. At 28 days after initial immunization, protective HAV antibody was produced in the serum of mice and the level of antibody was increased after boosting immunization. There was no significant difference in antibody level between the combination vaccine group and the corresponding monovalent vaccine group (P> 0.05) EV71 and HAV. After 28 days of initial immunization, the proportion of CD8 + cells in spleen of mice in each group was higher than that in CD4 + cells (P <0.001), and on the 7th day after immunization (P <0.001) , HAV full virus and EV71 polypeptide could not stimulate the spleen cells of each group to produce a positive reaction. At 7 days after boosting, the level of IL-4 secreted by spleen cells in each group was higher than that of IFNγ (P <0.05). Conclusions A combination of HAV and EV71 vaccine can induce good humoral immune response and induce cellular immune response in mice, but it is relatively weak. No mutual interference or synergism between the two antigens was found.