细粒棘球蚴亲肌肉抗原在小鼠中的免疫保护力研究

目的 对细粒棘球蚴亲肌肉抗原(myophilin antigen)进行原核表达、纯化并用小鼠实验评价其免疫学特性. 方法 提取总RNA,构建克隆质粒Eg myophilin/pGEM-T,然后将亲肌肉抗原基因亚克隆于表达载体pET28a,进行诱导表达;纯化重组蛋白并免疫小鼠.利用Western blot、ELISA鉴定其免疫学特性,并通过包囊计数,评价其免疫保护力. 结果 成功构建原核重组表达载体Eg myophilin/pET28a,并表达、纯化出浓度较高的亲肌肉抗原.ELISA检测显示,用纯化的亲肌肉抗原免疫小鼠,诱导产生了特异性抗体;Western blot鉴定该抗体能识别重组抗原及原头蚴.攻击感染实验表明亲肌肉抗原免疫小鼠的包囊数为0.93±2.1,对照组为7.13±10.21,两者差异有统计学意义,获得的保护力约为94.4%. 结论 成功表达细粒棘球蚴亲肌肉抗原,纯化后的重组蛋白具有抗原性和免疫原性,有望作为棘球蚴病疫苗候选分子.

Protective immunity of myophilin antigen of Echinococcus granulosus in mice

Objective To clone and express the gene encoding myophilin antigen of Echinococcus granulosus(Eg), and evaluate its immunogenicity and protective immunity in mice. Methods Total RNA was extracted from protoscoleces of the liver of human. The Eg myophilin gene was cloned into pGEM-T vector and transformed into Escherichia coli JM109. Then Eg myophilin gene was subcloned into pET28a and expressed. ICR mice were immunized with purified recombinant Eg myophilin (rEg myophilin), the specific sera were tested by Western blot and ELISA. Protection and immunogenicity of recombinant protein in mice were assessed by the rate of reduced larval weight. Results Eg myophilin was successfully produced in a recombinant E. coli expression system. The purified rEg myophilin and native antigen protoscoleces could be recognized by specific sera of mice immunized with rEg myophilin. ELISA detection showed the IgG level of sera of the rEg myophilin immunized group was higher than that of control group (P<0.05). The average number of hydatid cysts in rEg myophilin vaccinated mice and control group were 0.93±2.1,7.13±10.21, respectively(P< 0.05). The level of protective immunity was 94.4%. Conclusion The recombinant plasmid Eg myophilin/pET28a was successfully constructed and could be expressed in E. coli with antigenicity and immunogenicity, rEg myophilin could be a promising candidate for an effective vaccine to prevent echinococcosis.