摘要
目的 建立测定A、C、Y、W135群脑膜炎球菌(meningococcus,Men)多糖疫苗(groups A,C,Y,W135 menignococcal polysaccharide vaccine,MPV4)多糖含量的火箭免疫电泳(rocket immunoelectro-phoresis,RIE)法.方法 分别用A、C、Y、W135群Men菌液,A、C、Y、W135群Men菌液加弗氏佐剂,A、C、Y、W135群Men多糖-牛白蛋白结合物免疫家兔,制备特异性抗血清.将制备的抗血清以一定的比例加入琼脂糖凝胶制成平板,并将纯化的多糖作为定量参考品加入抗原孔进行RIE.以多糖含量和对应的RIE峰高作标准曲线并建立直线回归方程.采用优化的RIE法测定MPV4多糖含量和分子大小.结果 菌液加佐剂制备的抗血清效价较理想.在确定的电泳条件下,建立的RIE法制备的标准曲线呈现良好的线性关系,相关系数值均〉0.98.该法特异性较好,未检出各多糖间的交叉反应.采用该法测定的3批MPV4的多糖含量、分子大小及回收率均与先前的检定结果相符,均符合质控标准.结论 建立的RIE法可作为MPV4多糖抗原含量的测定方法.
Objective To establish a rocket immunoelectrophoresis (ME) method for determination of polysaccharide contents in groups A,C,Y,W135 meningococcal polysaccharide vaccine (MPV4). Methods Rabbits were immunized with meningococcal groups A,C,Y,W135 suspensions, mixture of Freund adjuvant with meningococcal suspension and meningococcal polysaccharide-bovine albumin conjugate, respectively.Specific antisera were prepared. The specific antisera were added to agarose gel with a certain proportion to pave plate, and pure polysaccharides as quantitative references were added into gel holes to carry out RIE.Standard curves were made with polysaccharide concentrations and precipitation peaks formed by RIE, and linear regression equations were established. Polysaccharide contents and molecular sizes in MPV4 were determined by the established RIE methed. Results Optimal antiserum titers were obtained by immunization with the mixture of Freund adjuvant and meningococcal suspension. In the definitive electrophoresis conditions,standard curves made by RIE method showed better linear correlation, and r values were all 〉0.98. Specificity of the RIE method was better, and no cross reactions among different polysaccharides were detected. The results of polysaccharide contents, molecular sizes and recovery rates of three batches of MPV4 determined by the RIE method were in accordance with those of previous determination and the quality control standards.Conclusion The established RIE method can be used as a determination method for polysaccharide contents of MPV4.
出处
《国际生物制品学杂志》
CAS
2011年第4期175-179,共5页
International Journal of Biologicals
关键词
免疫电泳
脑膜炎双球菌菌苗
多糖类
细菌
方法
Immunoelectrophoresis
Meningococcal vaccines
Polysaccharides,bacterial
Methods