PNEUMOCOCCAL POLYSACCHARIDE PROTEIN CONJUGATE VACCINES

肺炎球菌多糖蛋白结合疫苗

• 批准号: 6413390
• 负责人: CHI-JEN LEE
• 金额: --
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6413390
• 关键词: Streptococcus pneumoniae vaccine; bacterial polysaccharides; bacterial proteins; immune complex; immunoprecipitation; trypsin; vaccine development

The recently licensed pneumococcal conjugate vaccine contains 7 different types (4, 6B, 9V, 14, 18C, 19F, and 23F) conjugated to CRM197 protein. The immunochemical assay by rate nephelometry is being used for evaluation of individual PS content in polyvalent pneumococcal vaccines. Nephelometry measures the intensity of scattered light as it is passed through the antigen-antibody complex formed by the precipitin reaction. For quantitation of PS content by nephelometry, the optimum concentrations for PSs and antibodies need to be determined. The objective of the present study was therefore to determine the optimum conditions for quantitative determination of individual PS in a polyvalent PS-protein conjugate vaccine. A multivalent conjugate vaccine was treated with trypsin to digest the carrier protein, thus releasing PSs into solution. For reference, monovalent conjugate vaccines, 10 ug/ml, were treated with trypsin and the released PSs diluted to different concentrations.(0.5 to 10 ug/ml) The antisera used were type specific hyperimmune rabbit sera against each pneumococcal type. Standard curves for types 4, 6B, 9V, 14, 18C, 19F, and 23F PSs were proportional to the increase of PS concentration in the range of 0.5 to 10 ug/ml. An excess or high PS concentration resulted in inhibition of the precipitin reaction and a reduction in the Ag-Ab complexes and thereby a decreased scattering rate. The optimum concentration for these 7 PS types, displaying linearity when used with their respective antisera, was found to be 1-5 ug/ml. Rate nephelometry is simple in operation, utilizes small quantities of antigen and antibody, and yields reproducible and accurate data for the quantitative analyses of individual PSs in polyvalent vaccines. However, it requires the reference standards for PS antigens and specific high titered antisera. Substances and conditions that may interfere with the nephelometric immunoassay system are being examined.

最近获得许可的肺炎球菌结合疫苗包含7种不同的类型(4、6B、9V、14、18C、19F和23F)与CRM197蛋白结合。速率散射比浊法的免疫化学测定法正被用于评估多价肺炎球菌疫苗中的个体PS含量。散射比浊法测量散射光穿过沉淀反应形成的抗原-抗体复合体时的强度。为了用散射比浊法测定PS含量，需要确定PSS和抗体的最佳浓度。因此，本研究的目的是确定定量测定多价PS蛋白结合疫苗中单个PS的最佳条件。用胰酶处理多价结合疫苗，消化载体蛋白，从而释放PSS进入溶液。以10ug/ml的单价结合疫苗为参照，用胰酶处理释放的PSS，并将其稀释成不同浓度(0.5~10ug/ml)，所用抗血清为针对各肺炎球菌类型的型特异性高免兔血清。 4型、6B型、9V型、14型、18C型、19F型和23F型PSS的标准曲线与PS浓度在0.5~10ug/ml范围内呈线性关系，PS浓度过高或过高会抑制沉淀反应，减少Ag-Ab络合物，从而降低散射率。这7种PS的最适浓度为1-5微克/毫升，与各自的抗血清呈线性关系。 速率散射比浊法操作简单，使用少量的抗原和抗体，为多价疫苗中个体PSs的定量分析提供了可重复性和准确性的数据。然而，它需要PS抗原和特异性高滴度抗血清的参考标准。正在检查可能干扰散射比浊法免疫分析系统的物质和条件。