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目的优化评价肺炎链球菌多糖结合疫苗效力的调理吞噬试验(Opsonophagocytic assay,OPA)的条件。方法参考美国阿拉巴马大学的WHO指定实验室于2007年1月发布的A.02版实验规程,优化OPA的试验条件。分别以抗性和非抗性肺炎链球菌菌株作为靶细菌检测其对应型结合物受试血清的调理吞噬效价后,再与相应的ELISA效价比较,分析二者的相关性。结果 0.8%二甲基甲酰胺(DMF)诱导HL-60细胞分化4d后,细胞表面标志表达量分别为:CD11b和CD35≥55%、CD71≤15%,活细胞比例≥65%;各型肺炎链球菌工作种子批的复活率≥80%;补体的非特异性杀伤率为18%;各血清型均表现出良好的杀菌活性,且OPA法与ELISA法测定的血清抗体效价比较,相关系数和斜率非常接近。结论优化的OPA条件符合该试验要求,可用于肺炎链球菌荚膜多糖结合疫苗的效力评价。
Objective To optimize the conditions for Opsonophagocytic assay (OPA) for evaluating the efficacy of S. pneumoniae polysaccharide conjugate vaccine. Methods Reference Alabama University, the WHO designated laboratory in January 2007 released A.02 version of the experimental procedures, OPA optimization of the experimental conditions. Respectively, the resistant and non-resistant strains of Streptococcus pneumoniae were used as target bacteria to test the opsonophagocytic potency of the corresponding conjugate test sera, and then compared with the corresponding ELISA titer to analyze the correlation between the two. Results After HL-60 cells were differentiated for 4 days with 0.8% dimethylformamide (DMF), the expression of cell surface markers were: CD11b and CD35≥55%, CD71≤15% and live cells≥65% The resurrection rate of Streptococcus seed batch was ≥80%, the nonspecific killing rate of complement was 18%, and all serotypes showed good bactericidal activity. And the serum antibody titer determined by OPA method and ELISA method, the correlation coefficient and The slope is very close. Conclusion The optimized OPA conditions are in accordance with the requirements of this test and can be used to evaluate the efficacy of Streptococcus pneumoniae capsular polysaccharide conjugate vaccine.