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

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.

Abstract：

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.

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

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.