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目的初步评价肠道病毒71型(enterovirus 71,EV71)与乙型肝炎(简称乙肝)病毒(hepatitis B,HBV)联合疫苗中两种抗原之间的相互作用及其免疫小鼠诱导的特异性免疫应答水平。方法将EV71灭活疫苗抗原和重组HBV疫苗抗原(HBs Ag)按常规剂量,采用直接吸附法吸附Al(OH)3佐剂,配制成EV71-HBV联合疫苗,并设单苗组、佐剂对照组和空白对照组(PBS),分别于0、28 d各经腓肠肌免疫BALB/c小鼠1次,于初免后第2、4周和第2剂免疫后第2、4、8周经尾动脉采血,分离血清,采用双抗体夹心ELISA法检测血清中Anti-HBs水平,微量中和试验检测血清中EV71中和抗体效价;并于初免后第3、4周和第2剂免疫后第1、2、4周取小鼠脾脏,分离脾单个核细胞(mononuclear cell,MNC),采用酶联免疫斑点试验(enzyme-linked immunospot assay,ELISPOT)检测MNC中抗原特异性IFNγ的分泌水平。结果第2剂免疫后第2周,各疫苗组小鼠血清EV71中和抗体和Anti-HBs阳转率均达100%,各检测时间点疫苗组间抗体效价差异均无统计学意义(P>0.05),佐剂对照组和空白对照组抗体均无阳转。各疫苗组分泌IFNγ的斑点形成细胞(spot forming cell,SFC)数在5个检测时间点有所波动,第2剂免疫后第1周,所有疫苗组IFNγ分泌水平均明显升高,达5个检测时间点的峰值,第2剂免疫后第2周显著下降,但在第2剂免疫后第4周又小幅升高;除第2剂免疫后第2周EV71-HBV联合疫苗组SFC数显著高于EV71单苗组外(P<0.05),其他检测时间点EV71-HBV联合疫苗组与单苗组的SFC数差异均无统计学意义(P>0.05);佐剂对照组和空白对照组的ELISPOT检测结果均为阴性。结论一定剂量的EV71和HBV联合疫苗可诱导小鼠产生良好的体液免疫和细胞免疫应答,两种抗原间未发现相互作用。
OBJECTIVE: To evaluate the interaction between two antigens in enterovirus 71 (EV71) and hepatitis B (HBV) combination vaccine and the specific immunity induced by immunized mice Response level. Methods The EV71-HBV combination vaccine was prepared by adsorbing EV71 inactivated vaccine antigen and recombinant HBV vaccine antigen (HBsAg) by conventional adsorption method using direct adsorption method. The single vaccine and adjuvant control Group and blank control group (PBS). BALB / c mice were immunized with gastrocnemius muscle respectively on days 0 and 28 for 1, 2, 4 weeks after immunization and 2, 4, 8 weeks after immunization Serum anti-HBs levels were measured by double antibody sandwich ELISA. Serum levels of EV71 neutralizing antibody were detected by micro-neutralization assay. In the third and fourth week after immunization, the second immunization Spleen was collected at the 1st, 2nd, 4th and 4th week after spleen mononuclear cells (MNC) were isolated. The secretion of antigen-specific IFNγ was detected by ELISPOT . Results At the second week after the second immunization, the positive rates of serum EV71 neutralizing antibody and anti-HBs in the vaccine groups were both 100%. There was no significant difference in antibody titer between the two vaccine groups (P > 0.05), adjuvant control group and blank control group antibodies were no positive. The numbers of spot forming cells (SFCs) secreting IFNγ in each vaccine group fluctuated at 5 test points. The IFNγ secretion levels in all vaccine groups were significantly increased at the first week after the second immunization, reaching 5 The peak value at the time point of detection was significantly decreased at the second week after the second immunization, but slightly increased at the fourth week after the second immunization. In addition to the second immunization after the second immunization, the number of SFCs in the EV71-HBV combination vaccine group was significantly higher (P <0.05). There was no significant difference in the number of SFC between the EV71-HBV combination vaccine group and the single vaccine group at the other time points (P> 0.05). The adjuvant control group and blank control group ELISPOT test results were negative. Conclusion A certain dose of combination vaccine of EV71 and HBV can induce good humoral and cellular immune responses in mice, and no interaction is found between the two antigens.