脑膜炎球菌结合疫苗中细菌内毒素检查方法的建立及系统验证  

Establishment and verification of a method for testing bacterial endotoxin in meningococcal conjugate vaccines

作　　者：吴兵[1] 刘方蕾[1] 宣俊文[1] 于旭博 王轶峰[1] 孔素娟[1] 袁菲[1] 谢贵林[1] 赵志强[1] 

机构地区：[1]兰州生物制品研究所有限责任公司,兰州730046

出　　处：《中国新药杂志》2013年第24期2878-2887,2908,共11页Chinese Journal of New Drugs

基　　金：国家"重大新药创制"科技重大专项(2013ZX09402-302-215);甘肃省科技重大专项(0801NKDA003);甘肃省"十二五"陇药产业发展规划项目

摘　　要：目的:建立用于脑膜炎球菌结合疫苗中细菌内毒素含量的检测方法,研究其在脑膜炎球菌结合疫苗不同生产阶段应用的可行性,并对方法进行系统验证,以确保检查结果的真实性和准确性。方法:参考《中华人民共和国药典》2010年版三部附录,建立动态浊度法测定细菌内毒素含量,并验证生产过程中相关原辅材料本身对方法可靠性的影响,参考相关指导原则完成方法的系统验证。结果:方法的线性符合验证要求,测定范围为0.03～10 EU·mL-1;相关原辅料中的细菌内毒素含量均低于0.01 EU·mL-1,方法的专属性符合验证要求;添加实验结果显示标准品回收率均在80%～130%,表明方法准确性良好,样品本身对方法检定无影响;方法精密度和耐用性验证实验结果均能达到方法验证要求;方法定量限度为0.03 EU·mL-1。多次检查疫苗原液和成品多批次结果差异较小,细菌内毒素含量检定准确可靠。结论:经方法验证显示方法的线性、特异性、灵敏度和准确性等技术参数达到了相关验证要求,结果稳定可靠;各种原辅材料对细菌内毒素检查无影响。本研究为该疫苗细菌内毒素检查提供了更为快捷有效的质量控制手段,验证结果为进一步方法再验证提供了数据和应用支持。Objective： To establish a method （kinetic turbidimetric assay） for testing bacterial endotoxin in the quality control of meningococcal conjugate vaccines and evaluate its feasibility during the process of meningococcal conjugate vaccine preparation, and systematically verify the validation in the quality control for guaranteeing the accuracy and reality of the determination results. Methods： Kinetic turbidimetric assay included in Chinese Pharmacopoeia Volume III （2010 Edition） was referred to detect bacterial endotoxin contents, and verified in vaccine research according to the General Principles on Verification for Quality Control Methods issued by SFDA. The method reliability in bacterial endotoxin tests was subsequently verified by assessing the influence of raw and auxiliary materials used in vaccine manufacture. Results： Linearity of the method fulfilled the requirements, and the range of method determination was 0.03-10 EU·mL-1. Also all the bacterial endotoxin contents in the raw and auxiliary materials or reagents used in meningococcal conjugate vaccine manufacture were lower than 0.01 EU·mL-1, indicating that the selectivity （i.e. specificity） of the method met the requirements. The reference recovery was 80%-130% in the test of meningococcal conjugate vaccines added bacterial endotoxin, so accuracy of the method was relatively good. The precision and robustness met the requirements on verification of the methodology, and the limit of quantitation was 0.03 EU？mL-1 tentatively in the bacterial endotoxin test. Endotoxin content results of samples showed satisfactory reliability and accuracy because the differences of bacterial endotoxin contents in different batches of meningococcal conjugate vaccines were minimal among different bacterial endotoxin tests. Conclusion： The validation results indicate that the indexes of bacterial endotoxin test method including linearity, selectivity, accuracy, and sensitivity meet the requirements. The raw and auxiliary materials or lyophilized pro

关 键 词：细菌内毒素检查 脑膜炎球菌 结合疫苗 方法建立 方法验证 

分 类 号：R979.5[医药卫生—药品]