Poster Presentation Lancefield International Symposium for Streptococci and Streptococcal Diseases 2025

Development of a 10-plex electrochemiluminescent assay for quantifying specific serological immunoglobulin isotype and subclass responses to Streptococcus pyogenes antigens  (#213)

Michael Morici 1 2 3 , Jill Gilmour 1 4 , Hannah Frost 1 5 , Joshua Osowicki 5 , Andrew Steer 1 5 , Jonathan Carapetis 1 2 3 6 7 , Alma Fulurija 1 2 3 6
  1. The Australian Strep A Vaccine Initiative, Nedlands, Western Australia, Australia
  2. The Kids Research Institute Australia, Nedlands, Western Australia, Australia
  3. Wesfarmers Centre for Vaccines and Infectious Diseases, Nedlands, Western Australia, Australia
  4. JWG Global Lab Solutions, Dumfries, Scotland, UK
  5. Murdoch Children’s Research Institute, Parkville, Victoria, Australia
  6. The University of Western Australia, Nedlands, Western Australia, Australia
  7. Perth Children’s Hospital, Nedlands, Western Australia, Australia

Background 
The global burden of Streptococcus pyogenes is a major unmet public health challenge.  Development of a safe and effective vaccine requires standardized immune assays to generate reproducible and comparable antigen-specific serological data. Future trials including young children will require assays with high sensitivity, scalability, reproducibility, and precision, using only small sample volumes. 

Methods 
We developed 10-plex electrochemiluminescent-based assays using the Meso Scale Discovery (MSD) platform to quantify IgG, IgG subclasses, and IgA recognizing the S. pyogenes antigens streptolysin O (SLO), DNase B, SpyCEP, SpyAD, ScpA, Group A Carbohydrate (GAC), SpnA, Spy0843, OppA, and PulA. Assay parameters were optimized, specificity, dilutional linearity, and limits of quantitation were evaluated, and assay performance was assessed using samples from healthy donors. 

Results 
The assays demonstrated excellent specificity, sensitivity, dilutional linearity, and wide dynamic ranges for IgG, IgG subclasses and IgA detection. Using IVIG as a standard for IgG and pooled serum for IgA, standard curves were constructed and values in arbitrary units (AU)/mL were assigned, allowing sample concentrations to be interpolated from standard curves.  

Conclusion 
The 10-plex electrochemiluminescence assay provides a robust, flexible platform that builds on the previously reported 6-plex and adds detection options for IgA and IgG subclasses, making it ideal for seroprevalence studies, natural infection and human challenge studies, and advancing understanding of humoral immune responses to S. pyogenes antigens.