双蛋白载体A、C群脑膜炎球菌多糖结合物的免疫效果观察  

The immunological effect of group A and C meningococcal conjugate vaccine with double protein carriers

作　　者：李启明[1] 马智静[1] 张靖[1] 靳玉琴[1] 陈实[1] 张学峰[1] 刘梅影[1] 

机构地区：[1]北京生物制品研究所基因工程研究室,100024

出　　处：《中华微生物学和免疫学杂志》2011年第2期146-149,共4页Chinese Journal of Microbiology and Immunology

基　　金：国家十一五科技支撑计划资助项目（2008BA166803）

摘　　要：目的 比较和评价在A、C群脑膜炎球菌多糖结合物中应用两种蛋白作为载体的免疫效果.方法 构建不耐热肠毒素B亚单位（LTB）的克隆载体和表达载体.确定rLTB主要为可溶性表达.应用一步阳离子交换层析对rLTB进行纯化,获得较纯的目的蛋白.利用GM1-ELISA和非变性SDS-PAGE的方法确定纯化后的rLTB能够形成具有生物学活性的五聚体.最后利用化学方法（ADH方法）将通过基因工程手段获得的重组LTB五聚体蛋白与C群脑膜炎球菌多糖（GCMP）耦联,获得多糖蛋白结合物GCMP-rLTB.以单一蛋白破伤风类毒素（TT）为载体的A＋C群脑膜炎球菌多糖蛋白结合物（GAMP-TT和GCMP-TT）及同时应用两种蛋白载体的A＋C群脑膜炎球菌多糖蛋白结合物（GAMP-TT和GCMP-rLTB）分别通过腹腔注射途径免疫小鼠.应用ELISA方法分别检测小鼠血清中IgG水平.结果 单独以TT为载体的A＋C群脑膜炎球菌多糖蛋白结合物（GAMP-TT和GC-MP-TT）和同时以两种蛋白作为载体的A＋C群脑膜炎球菌多糖蛋白结合物（GAMP-TT和GCMP-rLTB）通过注射途径免疫小鼠,后者产生的血清多糖特异性IgG水平显著高于前者.结论 双蛋白载体在改善A、C群脑膜炎球菌多糖结合物疫苗的免疫原性方面具有明显优势.Objective To evaluate the immunogenicity of group A and C meningococcal polysaccharides conjugates using different proteins as carriers. Methods Heat-labile enterotoxin B subunit （LTB）pentamer form was expressed in E. coli. The target protein was identified and purified by cation-exchange chromatography. Then biological activity of rLTB was tested using GM1-ELISA. GCMP was conjugated to rLTB with the chemical method （ADH）. Furthermore, the mice were immunized with GAMp-TT/GCMP-TT conjugates and GAMP-TT/GCMP-rLTB conjugates via peritoneal. Finally the anti-polysaccharide antibody was detected. Results The GAMP-TT/GCMP-rLTB conjugate elicits remarkably higher serum antibodies in mice than GAMP-TT/GCMP-TT conjugate. Conclusion These results indicated that polysaccharide conjugates using different proteins as carriers were superior to those using only one protein as carrier.

关 键 词：重组大肠杆菌不耐热肠毒素B亚单位(rLTB) 多糖结合疫苗 A、C群脑膜炎球菌多糖 

分 类 号：R392[医药卫生—免疫学]