STANDARDIZATION OF AN OPSONOPHAGOCYTIC ASSAY FOR THE MEASUREMENT OF FUNCTIONAL ANTIBODY-ACTIVITY AGAINST STREPTOCOCCUS-PNEUMONIAE USING DIFFERENTIATED HL-60 CELLS

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 ser um samples were analyzed for the presence of functional antibody again st seven pneumococcal serogroups or serotypes (4, 6B, 9V, 14, 18C, 19F , and 23F) by using differentiated HL-60 cells (granulocytes) and peri pheral blood leukocytes (PBLs). Six of the 55 serum samples were from unvaccinated adult volunteers, 31 serum samples were from adults who r eceived one dose of the 14-valent or the 23-valent polysaccharide vacc ine, and 18 serum samples were from 16-month-old infants who received four doses of an investigational 7-valent polysaccharide-protein conju gate vaccine. The results of an opsonophagocytic assay,vith HL-60 cell s correlated highly with those of an assay with PBLs as effector cells (median r for seven serotypes = 0.87; P < 0.01). Opsonophagocytic tit ers were compared with the immunoglobulin G antibody concentrations de termined by enzyme-linked immunosorbent assay (ELISA). The r values fo r 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, a nd 0.62, respectively, when PBLs were used. The assay requires small a mounts of serum (40 mu l per serotype), making this test suitable for assaying infant sera. Culturable cells aid in assay standardization an d likely reduce donor-to-donor variability. This standardized assay, i n combination with the standardized ELISA, can be used to evaluate cur rent and developing pneumococcal vaccines, in which functional opsonop hagocytic antibody activity may correlate with protection against pneu mococcal disease.