Streptococcal serological tests provide evidence of prior infection by Group A Streptococcus (GAS). These tests are crucial to the diagnosis of the post-infectious immune sequelae acute rheumatic fever (ARF) and glomerulonephritis since these syndromes develop several weeks after a GAS infection. Current tests measure anti-streptolysin-O (ASO) and anti-DNaseB (ADB) antibodies. Though widely used, these tests have limitations including a slow rate of decay in antibody response that can increase the risk of false positives in settings where GAS is endemic, and incompatible methodology, requiring the two assays to be run in parallel. In this study, the utility of a novel GAS antigen, SpnA, was assessed in a multiplex bead-based assay that also incorporated streptolysin-O and DNaseB. Recombinant streptolysin-O, DNAseB and SpnA were conjugated to polystyrene beads for measurement of serum antibody binding in a Cytometric Bead Array. Multiplex assay were run on sera samples collected from participants in three groups: ARF; ethnically matched healthy children; and healthy adults. The ability of the antigens to detect a previous GAS exposure for ARF diagnosis was assessed using the 80th centile of the healthy children group as a cut-off (upper limit of normal). Using these experimentally determined cut‑offs SpnA had the highest sensitivity at 88% compared to 75% for ASO and 56% for DNaseB. In conclusion, SpnA has favourable immunokinetics for streptococcal serology, and the combination of SpnA with ASO and ADB in a multiplex assay should improve the efficiency and accuracy of streptococcal serology.