无细胞百日咳疫苗血清学效价检测方法的建立及适用性的初步验证

目的建立无细胞百日咳疫苗血清学效价检测方法(PSPT),并对其适用性进行初步验证。方法采用纯化的百日咳毒素(PT)和丝状血凝素(FHA)包被酶标板,以羊抗鼠IgG酶标抗体为指示抗体,建立检测小鼠抗PT抗体和抗FHA抗体的间接ELISA法,并对该方法进行初步验证。在此基础上建立PSPT法,并与MICA法进行相关性分析。结果建立了小鼠抗PT抗体ELISA和抗FHA抗体ELISA,标准曲线线性范围内R≥0.99;经验证,两系统与无关抗体之间均无交叉反应;检测限度分别为93.7500和59.0625mEU/ml,检测限量分别为100和70mEU/ml。将10批无细胞百日咳原液分别以PSPT法和MICA法进行效价测定,两者呈正相关,两种方法检测结果差异无统计学意义,且PSPT法比MICA法重复性更好。结论抗PT抗体ELISA和抗FHA抗体ELISA灵敏度高,准确性及精密性均较好,在此基础上建立起来的PSPT法有望替代MICA法,用于检测无细胞百日咳疫苗的效价。

Establishment and Adaptivity of Pertussis Serological Potency Test

Objective To establish pertussis serological potency test(PSPT)and verify its adaptivity. Methods Indirect ELISA for determination of murine anti-pertussis toxin(PT)and anti-filamentous hemagglutinin(FHA) antibodies(Abs) were established by coating a microtiter plate with purified PT and FHA respectively and serving enzyme-labeled sheep-anti-mouse IgG as indicating antibody then verified, based on which PSPT was established and analyzed for its relationship to modified intracerebral challenge assay(MICA). Results The indirect ELISA methods for murine anti-PT and anti-FHA Abs were established, with R values of not less than 0. 99 within the linear ranges of standard curves. Both the ELISA systems for anti-PT and anti-FHA Abs showed no cross reactions with irrelevant antibodies, and their detection limits were 93. 750 0 and 59. 062 5 mEU / ml, while the quantification limits were 100 and 70 mEU / ml, respectively. The potencies of 10 batches of bulks of acellular pertussis vaccine were determined by PSPT and MICA respectively, and the results showed no significant difference. The determination results by PSPT were positively related to those by MICA. Compared with MICA, PSPT showed good reproducibility. Conclusion The ELISA methods for anti-PT and anti-FHA Abs showed high sensitivity, accuracy and precision, and the PSPT established on the basis of the two ELISA methods might be used instead of MICA for determination of the potency of acellular pertussis vaccine.