Abstract
Group B Streptococcus (GBS) infections constitute a major cause of invasive disease during the first three months of life and an unmet medical need that could be addressed by maternal vaccination. The GBS capsular polysaccharides (CPSs) have shown promise as vaccine targets in clinical studies. A highly specific serological assay to quantify maternal and neonatal anti-CPS antibody levels will be instrumental for GBS vaccine licensure. Here, we describe the development and comparison of two novel multiplex immunoassays (MIAs) based on the Luminex technology for the quantification of IgG antibodies recognizing the five most frequent GBS capsular variants (Ia, Ib, II, III, and V) out of the ten types identified. The first assay is based on the use of biotinylated CPSs coupled to streptavidin-derivatized magnetic microspheres (Biotin-CPS MIA), while the second is a sandwich assay with plain CPSs coupled to magnetic microspheres coated with polysaccharide-specific mouse monoclonal antibodies (Sandwich MIA). Both assays showed good specificity, linearity, and precision, although the Biotin-CPS MIA presented higher sensitivity and lower complexity than the Sandwich MIA. A panel of human sera representing a wide range of anti-CPS IgG concentrations was tested in parallel by the two assays, which resulted in comparable titers. Our data support the preservation of antigenic epitopes in the biotinylated polysaccharides and the suitability of the Biotin-CPS MIA for the precise determination of GBS anti-CPS IgG concentrations in human sera.IMPORTANCE Group B streptococcal infections can cause death in neonates up to 3 months of age. Intrapartum antibiotic prophylaxis in GBS-colonized mothers has limited early infections but has no impact after the first week of life. The development of a maternal vaccine to address this unmet medical need has been identified as a priority by the World Health Organization, and the GBS CPSs are considered the best antigen targets. However, to date there are no accepted standardized assays to measure immune responses to the investigational vaccines and for establishment of serocorrelates of protection. Here, we describe the performance of two microsphere-based pentaplex immunoassays for the determination of antibodies recognizing the five most frequent GBS serotypes. Our data confirm that an assay based on biotinylated polysaccharides coupled to streptavidin microspheres would be suitable for the intended purpose.
Highlights
Group B Streptococcus (GBS) infections constitute a major cause of invasive disease during the first three months of life and an unmet medical need that could be addressed by maternal vaccination
We evaluated the performance of two multiplex immunoassays (MIAs) developed in parallel based on the Luminex technology for quantification of IgG antibodies against capsular polysaccharides (CPSs) of the most prevalent GBS serotypes, i.e., Ia, Ib, II, III, and V
Pentaplex Assays for Anti-GBS Antibody Measurement assumed that a biotin/CPS molecular ratio around 1% (1 mol of biotin per 100 mol of CPS repeating units (RU)) would result in minimal impact on the epitope and a low residual amount of unreacted CPS
Summary
Group B Streptococcus (GBS) infections constitute a major cause of invasive disease during the first three months of life and an unmet medical need that could be addressed by maternal vaccination. The first assay is based on the use of biotinylated CPSs coupled to streptavidinderivatized magnetic microspheres (Biotin-CPS MIA), while the second is a sandwich assay with plain CPSs coupled to magnetic microspheres coated with polysaccharide-specific mouse monoclonal antibodies (Sandwich MIA). Clinical studies to assess GBS vaccine efficacy are challenging to design, as they would require a large number of participants; licensure of a GBS vaccine may depend on the establishment of immunological correlates of protection [20, 21] This implies that the development of a quality-ensured standardized serological assay for the precise quantification of antibody levels against the major GBS CPS serotypes is important for assessing immune responses in vaccinated subjects and for establishing antibody levels predictive of neonatal protection from casecontrol observational studies
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