目的：建立肺炎球菌疫苗免疫效果评价的功能性抗体滴度检测方法。方法参考美国阿拉巴马大学的多重调理吞噬实验(MOPA)方法，构建并鉴定HL-60效应细胞库、肺炎球菌工作菌种库和补体；在此基础上对5份血清样本进行调理吞噬活性检测。结果 HL-60细胞分化后表面标志表达量CD35为71.78%，CD71为10.97%，活细胞比例为72.96%。13个血清型的肺炎链球菌工作菌种的冻存复活率均高于90%；抗生素抗性与敏感性与预期结果一致；工作稀释度均≥1000。补体的非特异性杀菌率均≤70%。血清样本的杀菌曲线均为标准的S型，平行孔间的差异不超过3倍。结论效应细胞、工作菌种、补体及血清样本的检测结果均符合阿拉巴马大学参考方法的标准要求，成功建立了基于MOPA的功能性抗体检测方法。%Objective To establish the functional antibody detection method for evaluation of immune effect of pneu-mococcus vaccine. Methods The effector cell cultures, working bacteria stocks and complement, which were important components of multiplexed opsonophagocytic killing assay (MOPA) method were constructed and detected for applica-bility according to the reference method published by WHO reference laboratory in US University of Alabama at Birm-ingham (UAB). The opsonophagocytic titers of five sera samples were determined using the established MOPA method. Results After differentiation of HL-60 cells, the expression of cell surface markers CD35 and CD71 were 71.78% and 10.97%, respectively, and the proportion of live cells was 72.96%. The percentage recovery of working bacteria stocks of all serotypes were more than 90%. The antibiotic sensitivity and resistance of the frozen assay stocks were accor-dance with the expected results. The optimal dilution factor for all the assay stocks were more than 1000. The non-specific killing rates of complement for all serotypes were less than 70%. The sample's killing curves for all serotypes were sigmoid shape, and the opsonophagocytic titers of the duplicates were differ by less than 3-fold. Conclusion The detection results of the effector cell cultures, working bacteria stocks, complement and sera samples meet the require-ments of reference method of UAB, indicating the MOPA method is successfully established in this study.
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