以HBc颗粒为呈现载体的猪囊虫疫苗的构建及其免疫学研究

目的构建以HBc为载体的带有三个猪囊虫抗原表位(n1,n2,n3)的重组融合表达质粒pET-Δc-3n,表达出融合蛋白并纯化,通过免疫小鼠研究其体液免疫效果。方法用PCR法将三个猪囊虫表位分别插入HBc序列的第78-79位之间以及149位之后,再将该序列克隆入pET28a载体中,构建重组表达质粒,用大肠杆菌BL21作宿主菌表达出融合蛋白,并命名为PCCE,纯化后免疫小鼠,检测小鼠的体液免疫应答。用绦虫卵攻击免疫小鼠,并观察疫苗的保护作用。结果测序结果表明重组质粒构建成功,SDSPAGE显示融合蛋白表达正确,并有多聚体形成现象,ELISA检测到高滴度抗体。小鼠体内绦虫卵攻击试验表明疫苗PCCE的相对保护率为89%。结论以HBc为呈现载体的猪囊虫疫苗被成功表达和纯化,该疫苗能诱导较强的体液免疫反应,免疫小鼠对绦虫卵攻击具有较好的免疫保护作用。提示该疫苗PCCE可能具有预防囊虫病的潜在价值。

Construction of a vaccine against Taenia solium cysticercosis based the HBc particles as the expression vector

A recombinant plasmid pETΔc-3n was constructed in which 3 epitopes of Taenia solium cysticercus were inserted into the trunicated HBc vector.The fusion protein expressed was purified and used to immunize mice to induce the humoral immune responses.By PCR, two epitopes were inserted into the sequence of amino acid residues 78-79 of HBc. particles and one epitope was added to the C-terminus of HBc149.The PCR product was then cloned into pET28a to construct recombinant expression plasmid. And it was transformed to E.coli BL21 to express the fusion protein through induction. After purification, this protein termed as PCCE was used to immunize mice to induce humoral immune responses. The immunized mice were challenged with infectious cysticercus eggs and the relative protection rate was observed later on.It was found the recombinant plasmid was successfully construtcted as demonstrated by DNA sequencing analysis.A fusion protein with correct molecular mass was expressed and confirmed by SDS-PAGE with polymer formation. High antibody titres were elicited in mice immunized with PCCE, as demonstrated by ELISA,and the relative protection rate was 89% as demonstrated by the challenge test. It is concluded that a vaccine against cysticercosis based on the HBc particle vector containing 3 protective epitopes has been successfully prepared and purified.Its property to induce strong humoral immune responses and high protection rate in mice permits for its use as a potential means to prevent T.solium cysticercosis.