肠道病毒71型与乙型肝炎病毒联合疫苗免疫原性的初步评价

发布时间：2018-09-18 06:58

【摘要】：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(P0.05),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(P0.05),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(P0.05);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。
[Abstract]:Objective to evaluate the interaction between the two antigens of enterovirus 71 (enterovirus 71) and hepatitis B virus (hepatitis) hepatitis B virus (HBV) combined vaccine and the level of specific immune response induced by immunization in mice. Methods EV71 inactivated vaccine antigen and recombinant HBV vaccine antigen (HBs Ag) were prepared by direct adsorption method to adsorb Al (OH) 3 adjuvant. The adjuvant control group and the blank control group were immunized with gastrocnemius muscle once on day 028, respectively. The blood samples were collected from caudal artery at the 4th week after the first immunization and the 2nd week after the 2nd dose immunization. The serum samples were isolated and the serum Anti-HBs levels were detected by double antibody sandwich ELISA method. The neutralization antibody titers of EV71 in serum were detected by microneutralization test, and the spleen of mice was collected at the 3rd week and 2nd week after immunization. Splenic mononuclear cells (mononuclear cell,MNC) were isolated and the levels of antigen-specific IFN 纬 in MNC were detected by enzyme linked immunoblot assay (enzyme-linked immunospot assay,ELISPOT). Results at the second week after the second dose immunization, the serum EV71 neutralizing antibody and Anti-HBs positive conversion rate of each vaccine group were 100, and there was no significant difference in antibody titer between each test time point (P0.05), and there was no positive conversion between adjuvant control group and blank control group. The number of (spot forming cell,SFC of IFN 纬 secreted by dot-forming cells in each vaccine group fluctuated at 5 test time points, and the level of IFN 纬 secretion in all vaccine groups increased significantly at the first week after the second dose immunization, reaching the peak value at 5 test time points. At the 2nd week after the second dose immunization, it decreased significantly, but increased slightly at the 4th week after the second dose immunization. The number of SFC in EV71-HBV combined vaccine group was significantly higher than that in EV71 single vaccine group at the second week after immunization (P0.05), but there was no significant difference in SFC number between EV71-HBV combined vaccine group and single vaccine group at other time points (P0.05); the number of SFC in adjuvant control group and blank control group was significantly higher than that in control group (P0.05). The results of ELISPOT were negative. Conclusion A certain dose of EV71 and HBV combined vaccine can induce good humoral and cellular immune responses in mice. No interaction between the two antigens was found.
【作者单位】：中国医学科学院北京协和医学院医学生物学研究所生物制品四室;
【基金】：国家十二五“重大新药创制”科技重大专项(2015ZX09101-031)
【分类号】：R373.2;R392-33

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