1、3、4、5、6B、7F、9V、14、18C、19A、19F和23F型兔源肺炎球菌特异性血清国家参考品的制备  

Preparation of national references for serotypes 1,3,4,5,6B,7F,9V,14,18C,19A,19F and 23F pneumococcal specific serums from rabbit origin

作　　者：陈琼[1] 王珊珊[1] 石继春[1] 王春娥[1] 李红[1] 李茂光[1] 叶强[1] CHEN Qiong;WANG Shan-shan;SHI Ji-chun;WANG Chun-e;LI Hong;LI Mao-guang;YE Qiang(National Institutes for Food and Drug Control,Beijing 100050,China)

机构地区：[1]中国食品药品检定研究院卫生部生物技术产品检定方法及其标准化重点实验室,北京100050

出　　处：《中国生物制品学杂志》2022年第2期179-183,共5页Chinese Journal of Biologicals

基　　金：国家重大新药创制专项(2018ZX09738006-006).

摘　　要：目的制备1、3、4、5、6B、7F、9V、14、18C、19A、19F和23F型兔源肺炎球菌特异性血清国家参考品,用于速率比浊法检测疫苗中的多糖含量。方法采用双光径速率浊度免疫分析系统(IMMAGE 800)非竞争性浊度模式进行检测,多糖体积21μL,血清体积21μL,反应缓冲液体积300μL,增益系数为4,反应时间为3.5 min。确定1、3、4、5、6B、7F、9V、14、18C、19A、19F和23F型兔源肺炎球菌特异性血清候选参考品的工作浓度、线性范围、检测限、定量限,并验证专属性和稳定性。采用本实验制备的血清国家参考品及市售血清同时检测2个厂家的23价肺炎球菌多糖疫苗的多糖含量。结果确定各型血清候选参考品的工作浓度在原倍至4倍稀释度之间,在相应工作浓度下,多糖浓度在1~6μg/mL范围时,与响应值呈良好的线性关系,相关系数(R2)均>0.980;检测限和定量限均低于多糖候选参考品最低点(1μg/mL)的反应值;各型血清候选参考品特异性良好,无交叉反应;血清候选参考品于2~8℃保存28 d后仍具有良好的稳定性。除厂家2疫苗的6B和9V型外,2种血清对厂家2疫苗其他型别及厂家1疫苗所有型别多糖含量检测结果的相对标准偏差(RSD)<15%。结论成功制备了12种兔源肺炎球菌特异性血清国家参考品,可用于速率比浊法检测23价肺炎球菌多糖疫苗的多糖抗原含量。Objective To prepare the national references for serotypes 1,3,4,5,6B,7F,9V,14,18C,19A,19F and 23F pneumococcal specific serum from rabbit origin and use for the determination of polysaccharide content in vaccine by rate turbidimetry.Methods Double light path rate turbidity immunoassay system(IMMAGE 800)was adopted in non-competitive nephelometric mode.Both the volumes of polysaccharide and serum samples were 21μL,while that of reaction was 300μL,the gain efficient was 4,and the reaction time was 3.5 min to determine the working dilutions,linear ranges,limits of detection and quantitation,specificity and stability of candidate references for serotypes 1,3,4,5,6B,7F,9V,14,18C,19A,19F and 23F pneumococcal specific serum.The polysaccharide contents in 23-valent pneumococcal polysaccharide vaccines from two manufacturers were determined by the prepared national references and commercial serum separately.Results The working dilutions of candidate references of various serotypes were determined as 0~4,at which the linear range of polysaccharide was 1~6μg/mL,with R2 values of more than 0.980.Both the limits of detection and quantitation were lower than the lowest point of standard curves(1μg/mL).The candidate references of various serotypes showed high specificity without cross reactions,and good stability after storage at 2~8℃for 28 d.Except those of serotypes 6B and 9V from manufacturer 2,the relative standard deviation(RSD)of determination results of vaccine of other serotypes from manufacturer 2 and those of vaccines of all the serotypes from manufacturer 1 by the prepared national references and the commercial serum were less than 15%.Conclusion Twelve national references of pneumococcal specific serum from rabbit origin were sucessfully prepared,which may be used for determination of the polysaccharide antigen contents in 23-valent pneumococcal polysaccharide conjugate vaccines by rate turbidimetry.

关 键 词：肺炎球菌 特异性血清 国家参考品 速率比浊法 

分 类 号：R378.14[医药卫生—病原生物学] R392-33[医药卫生—基础医学]