No abstract

A human reference serum pool, lot 89-S, has been developed for use in quantitating concentrations of antibody to Streptococcus pneumoniae. Weight-based units have been assigned to antibodies to 11 pneumococcal polysaccharide (PnPs) serotypes (1, 3, 4, 5, 6B, 7F, 9V, 14, 18C, 19F, and 23F) by using enzyme-linked immunosorbent assay methodology and a human standard reference serum, USNRP IS 1644. The experimentally derived assignments for anti-PnPs antibodies of the immunoglobulin G (IgG), IgM, and IgA isotypes in lot 89-S correlate well to the separately determined immunoglobulin assignment. These assignments for this antipneumococcal standard serum were used to quantitate IgG, IgM, and IgA isotype levels and the total immunoglobulin level in pediatric samples from a pneumococcal conjugate vaccine trial. The data indicate that these assignments may be used to assess levels of antibody to PnPs serotypes in human serum.

Streptococcus pneumoniae (pneumococcus) polysaccharide serotype-specific antibodies that have opsonophagocytic activity are considered a primary mechanism of host defense against pneumococcal disease. In vitro opsonophagocytic assays (OPAs) with antibody and complement to mediate opsonophagocytic killing of bacteria have been designed and developed as an adjunct to the standardized serum immunoglobulin G antipneumococcal capsular polysaccharide enzyme immunoassay to assess the effectiveness of pneumococcal vaccines. OPA presents challenges for assay standardization and assay precision due to the multiple biologically active and labile components involved in the assay, including human polymorphonuclear leukocytes or cultured effector cells, bacteria, and complement. Control of these biologically labile components is critical for consistent assay performance. An approach to validating the performance of the assay in accordance with International Conference for Harmonization guidelines, including its specificity, intermediate precision, accuracy, linearity, and robustness, is presented. Furthermore, we established parameters for universal reagents and standardization of the use of these reagents to ensure the interlaboratory reproducibility and validation of new methodologies.