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目的建立速率比浊法检测13价肺炎球菌结合疫苗中的结合多糖抗原含量,并对方法进行验证。方法采用免疫化学系统(IMMAGE 800),选择非竞争性浊度模式,样品或标准品20μl,血清20μl,反应缓冲液200μl,增益系数为4,反应时间为2.5 min;样品离心沉淀后用缓冲溶液复溶,经Na OH解吸附法处理样品和标准品。对方法进行线性、重复性、准确性、专属性、适用性验证,并采用建立的方法检测3批13价肺炎球菌结合疫苗样品中的各型多糖抗原含量。结果在设定的条件下,各型多糖抗原浓度在1~6μg/ml范围内,线性良好(相关系数>0.99);各型多糖抗原含量重复检测的相对标准偏差(RSD)均低于8%;13型多糖结合抗原含量的回收率在70%~130%之间;6A与19A型特异性血清与其他12型多糖抗原有交叉反应,但与阳性对照的比值均较低(<5%),其他12种特异性血清特异性良好,无交叉反应;除1、3型抗原外,其余型别的游离多糖抗原基本不干扰结合抗原的检测结果。结论建立的速率比浊法检测13价肺炎球菌结合疫苗中的结合多糖抗原含量重复性、准确性良好,1、3型结合抗原不适用于直接离心沉淀获得。
Objective To establish a rate turbidimetric method to detect the content of binding polysaccharide antigen in 13-valent pneumococcal conjugate vaccine and validate the method. Methods Immune chemistry system (IMMAGE 800) was used to select non-competitive turbidimetric model. The sample or standard 20μl, serum 20μl, reaction buffer 200μl, gain coefficient 4 and reaction time 2.5 min. After the sample was centrifuged and precipitated with buffer solution Re-dissolved, NaOH desorption of samples and standards. The method was validated by linearity, repeatability, accuracy, specificity, applicability and the established method was used to detect the content of each polysaccharide antigen in three batches of 13-valent pneumococcal conjugate vaccine samples. Results Under the given conditions, the relative standard deviations (RSDs) of all polysaccharide antigen samples were all within the range of 1 ~ 6μg / ml with good linearity (correlation coefficient> 0.99). The relative standard deviations (RSDs) ; The recovery rate of type 13 polysaccharide binding antigen was between 70% and 130%; type 6A and 19A specific sera cross-reacted with other type 12 polysaccharide antigens, , Other 12 kinds of specific serum specific good, no cross-reaction; in addition to type 1, the other type of free polysaccharide antigen basically does not interfere with the binding antigen test results. Conclusion The established turbidimetric method can be used to detect the content of binding polysaccharide antigen in 13-valent pneumococcal conjugate vaccine with good repeatability and accuracy. The type 1 and type 3 antigen are not suitable for direct centrifugation.