PROTECTIVE IMMUNITY IN MICE TO PNEUMOCOCCAL POLYSACCHARIDE PROTEIN

改进肺炎球菌疫苗的一种方法是补充 含蛋白质抗原的多糖疫苗。肺炎溶血素是一种重要的 大多数肺炎球菌产生的毒力因子。因此，停用 肺炎溶血素可以作为载体蛋白与 肺炎球菌PS形成PS-蛋白质结合物。停用的 从突变型ply基因中克隆并表达肺炎溶血素。 大肠杆菌(类肺炎杆菌，PDB)，或用0.1%的19F肺炎溶血素治疗 甲醛在37℃下放置30-60分钟。该蛋白质被共价连接到 PS用硫醇和蛋白质用硫醇衍生化后 溴乙酰基。连接了9V PS或19F PS的连接物 在怀孕期间给小鼠注射灭活的溶血素 和/或哺乳，并在早期生命中进入后代。反PS 免疫幼鼠的抗肺炎溶血素效价显著高于对照组。 高于未免疫对照组。当幼小的老鼠 用9V型或19F型肺炎球菌攻击后，细菌被清除 免疫小鼠的血液比对照小鼠的血液更有效。 用19F细胞攻击的免疫小鼠的存活率为 也长于对照组。这些结果表明， 免疫幼鼠可诱导保护性免疫。 孕期、哺乳期或婴儿期早期的结合体。 为了研究分子大小对抗体反应的影响，19F PS 从PS到寡糖被裂解成不同的大小，然后 与灭活的19F溶气素偶联。19F-Ig G抗体反应 已经下定决心。使用大分子尺寸PS生产的偶联物 更高的血清总19F-IgG水平以及3.5-4.5倍的IgG1水平 幼鼠的IgG3抗体水平是对照组的2.5到16.8倍 使用较小的PS或低聚糖进行偶联。

One approach to improve the pneumococcal vaccine is to supplement the polysaccharide vaccine with protein antigens. Pneumolysin is an important virulence factor produced by most pneumococci. Thus, inactivated pneumolysin can act as a carrier protein to link covalently with the pneumococcal PS to form a PS-protein conjugate. The inactivated pneumolysin was prepared from the mutant ply gene cloned and expressed in E. coli (pneumolysoid, pdB), or treatment of 19F pneumolysin with 0.1% formaldehyde at 37 C for 30-60 min. The protein was covalently linked to PS after derivatization of the PS with a thiol group and the protein with a bromoacetyl group. The conjugates, in which 9V PS or 19F PS was linked to the inactivated pneumolysin were injected into mice during pregnancy and/or lactation, and into their offspring during early life. The anti-PS and anti-pneumolysin titers of the immunized young mice were significantly higher than those of non-immunized controls. When the young mice were challenged with type 9V or type 19F pneumococci the bacteria were cleared more effectively from the blood of immunized mice than from control mice. The survival rate for the immunized mice challenging with 19F cells was also longer than that for the control group. These results indicated that the protective immunity could be induced in young mice by immunization with the conjugate during gestation, lactation or early infancy. To study the effect of molecular size on the antibody response, type 19F PS was cleaved into different sizes from PS to oligosaccharide, then conjugated with inactivated 19F pneumolysin. The 19F IgG antibody response was determined. The conjugate that used large molecular size PS produced higher serum levels of total 19F IgG as well as 3.5 to 4.5 fold higher IgG1 and 2.5 to 16.8 fold higher IgG3 antibodies in young mice, than the conjugates using smaller PS or oligosaccharide.