产肠毒素大肠埃希菌K99-987P-F41融合蛋白在小鼠体内免疫效果的评价

Immunological efficacy in vivo of fusion protein K99-987P-F41 of enterotoxigenic Escherichia coli in mice

目的评价产肠毒素大肠埃希菌(enterotoxigenic Escherichia coli,ETEC)K99-987P-F41融合蛋白在小鼠体内的免疫效果。方法制备ETEC融合蛋白K99-987P-F41亚单位疫苗、ETEC全菌体灭活疫苗、ETEC天然菌毛K99-987P-F41混合蛋白亚单位疫苗及ETEC重组菌毛K99-987P-F41混合蛋白亚单位疫苗,分别免疫小鼠,同时以免疫PBS的小鼠作为对照。ELISA法检测小鼠血清中IgG、IL-4和IFNγ的水平及小鼠粪便和小肠冲洗液中分泌型免疫球蛋白A(secreted immunogobulin A,sIgA)的含量;MTT法检测小鼠脾淋巴细胞增殖情况,并进行攻毒试验。结果 ETEC融合蛋白K99-987P-F41重组亚单位疫苗组小鼠血清中的抗ETEC的特异性IgG抗体、IL-4、IFNγ、粪便和小肠冲洗液中的sIgA、小鼠脾脏B和T淋巴细胞增殖能力均显著高于PBS对照组(P<0.05);ETEC融合蛋白K99-987PF41重组亚单位疫苗对小鼠的保护率为80%,与其他3种疫苗的小鼠保护率相近,而PBS对照组为0。结论 ETEC融合蛋白K99-987P-F41可有效诱导小鼠产生较高滴度的特异性抗体,可作为ETEC重组亚单位疫苗候选蛋白。

Objective To evaluate the immunological efficacy in vivo of enterotoxigenic Escherichia coli（ETEC） fusion protein K99-987P-F41 in mice. Methods Fusion protein K99-987P-F41 subunit vaccine, inactivated whole cell ETEC vaccine, natural pili K99-987P-F41 hybrid protein subunit vaccine and recombinant pili K99-987P-F41 hybrid protein subunit vaccine were prepared and immunized to mice respectively, using PBS as control. The IgG, IL-4 and IFNγ levels in sera as well as sIgA contents in feces and intestine washes were determined by ELISA, while the spleen lymphocyte cell proliferation was measured by MTT assay, and challenge test was carried out. Results The specific antibody level against ETEC, IL-4, IFNγ and s Ig A levels, as well as B and T lymphocyte proliferation levels of mice immunized with fusion protein K99-987P-F41 subunit vaccine were significantly higher than those in PBS control group（P 〈 0. 05）. The protective rate of the vaccine was 80%, which was similar to those of the other three vaccines. However, the protective rate of PBS was zero. Conclusion Fusion protein K99-987P-F41 efficiently induced high antibody level in mice, which might be used as a candidate protein of ETEC subunit vaccine.