Standardization of an opsonophagocytic assay for the measurement of functional antibody activity against Streptococcus pneumoniae using differentiated HL-60 cells
Standardization of an opsonophagocytic assay for the measurement of functional antibody activity against Streptococcus pneumoniae using differentiated HL-60 cells
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DOI：
10.1128/cdli.4.4.415-422.1997
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发表时间：
1997-07-01
期刊：
影响因子：
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通讯作者：
Carlone, GM
中科院分区：
文献类型：
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作者：
RomeroSteiner, S;Libutti, D;Carlone, GM
宿主对肺炎球菌疾病的保护主要是通过吞噬作用介导的。我们开发并标准化了一种使用HL-60细胞（人早幼粒细胞白血病细胞）的调理吞噬试验。采用分化的HL-60细胞（粒细胞）和外周血白细胞（PBL）分析了55份血清样品中是否存在针对7种肺炎球菌血清群或血清型（4、6 B、9V、14、18 C、19 F和23 F）的功能性抗体。55份血清样本中有6份来自未接种疫苗的成年志愿者，31份血清样本来自接种1剂14价或23价多糖疫苗的成年人，18份血清样本来自接种4剂研究用7价多糖蛋白结合疫苗的16个月大婴儿。HL-60细胞的调理吞噬试验结果与PBL作为效应细胞的试验结果高度相关（7种血清型的中位数r = 0.87; P < 0.01）。调理吞噬滴度与酶联免疫吸附试验（ELISA）测定的免疫球蛋白G抗体浓度进行比较。当使用HL-60细胞作为效应细胞时，血清群或血清型4、6 B、9V、14、18 C、19 F和23 F的r值分别为0.61、0.60、0.67、0.90、0.61、0.39和0.57，当使用HL-60细胞作为效应细胞时，血清群或血清型4、6 B、9V、14、18 C、19 F和23 F的r值分别为0.56、0.47、0.61、0.90、0.71、0.31、0.32和0.33。而使用PBL时，分别为0.62和0.62。该试验需要少量血清（每种血清型40 μ l），因此该试验适用于测定婴儿血清。可培养细胞有助于测定标准化，并可能减少供体间的变异性。这种标准化的检测，结合标准化的ELISA，可用于评估目前和发展中的肺炎球菌疫苗，其中功能性调理吞噬抗体活性可能与对肺炎球菌疾病的保护相关。
Host protection against pneumococcal disease is primarily mediated by phagocytosis. We developed and standardized an opsonophagocytic assay using HL-60 cells (human promyelocytic leukemia cells). Fifty-five serum samples were analyzed for the presence of functional antibody against seven pneumococcal serogroups or serotypes (4, 6B, 9V, 14, 18C, 19F, and 23F) by using differentiated HL-60 cells (granulocytes) and peripheral blood leukocytes (PBLs). Six of the 55 serum samples were from unvaccinated adult volunteers, 31 serum samples were from adults who received one dose of the 14-valent or the 23-valent polysaccharide vaccine, and 18 serum samples were from 16-month-old infants who received four doses of an investigational 7-valent polysaccharide-protein conjugate vaccine. The results of an opsonophagocytic assay,vith HL-60 cells correlated highly with those of an assay with PBLs as effector cells (median r for seven serotypes = 0.87; P < 0.01). Opsonophagocytic titers were compared with the immunoglobulin G antibody concentrations determined by enzyme-linked immunosorbent assay (ELISA). The r values for serogroups or serotypes 4, 6B, 9V, 14, 18C, 19F, and 23F were 0.61, 0.60, 0.67, 0.90, 0.61, 0.39, and 0.57, respectively, when HL-60 cells were used as effector cells and 0.56, 0.47, 0.61, 0.90, 0.71, 0.31, and 0.62, respectively, when PBLs were used. The assay requires small amounts of serum (40 mu l per serotype), making this test suitable for assaying infant sera. Culturable cells aid in assay standardization and likely reduce donor-to-donor variability. This standardized assay, in combination with the standardized ELISA, can be used to evaluate current and developing pneumococcal vaccines, in which functional opsonophagocytic antibody activity may correlate with protection against pneumococcal disease.