Development and characterisation of a four-plex assay to measure Streptococcus pyogenes antigen-specific IgG in human sera

The measurement of antibodies to vaccine antigens is crucial for research towards a safe and effective vaccine for Streptococcus pyogenes (Strep A). We describe the establishment and detailed characterisation of a four-plex assay to measure IgG to the Strep A vaccine antigens SpyCEP, Slo, SpyAD and GAC using the Luminex multiplex platform. A standard curve was established and characterized to allow the quantification of antigen-specific IgG. Assay specificity, precision, linearity, reproducibility and repeatability were determined via the measurement of antigen-specific IgG from pooled human serum. The assay is highly specific, reproducible and performs well across a large range of antibody concentrations against all four antigens. It is, therefore, suitable for future clinical trials in humans with a four-component vaccine, as well as for seroepidemiological studies to gain insights into naturally occurring immunity.