Abstract
Neisseria meningitidis is a gram negative diplococcal bacterium. Worldwide, N. meningitidis is the leading cause of bacterial meningitis and sepsis, with five serogroups (A, B, C, Y, and W-135) responsible for the majority of the disease. Multivalent (A, C, Y, and W-135) polysaccharide and conjugate vaccines have been licensed in the United States and elsewhere and are widely available. We have developed a multi-plexed electrochemiluminescent assay to quantitate serum antibody responses to meningococcal polysaccharides A, C, Y, and W-135 to allow for rapid evaluation of li- censed and investigational vaccines. A 96-well plate containing a carbon electrode arrayed with polysaccharides A, C, Y, and W-135 on separate spots within each well has been developed for simultaneous detection of polysaccharidespecific antibodies in serum samples from vaccinated individuals. The assay conditions were optimized using the anti-meningococcal serogroup A/C reference serum pool, CDC 1992 (NIBSC 99/706), through evaluation of plate types, coating polysaccharide concentrations, and blocking and serum diluent buffers. Comparison of single and multiplex assays demonstrated the sensitivity, specificity, and speed of the multi-plex format for the quantification of serum antibody responses to N. meningitidis polysaccharides A, C, Y and W-135.
Highlights
We have developed a multi-plexed electrochemiluminescent assay to quantitate serum antibody responses to meningococcal polysaccharides A, C, Y, and W-135 to allow for rapid evaluation of licensed and investigational vaccines
Infections caused by the bacterium Neisseria meningitisdis remain an important medical need, in the developing world where periodic outbreaks are responsible for thousands of deaths [1]
The impact of various coating concentrations (5, 50, and 250 ng/well) of meningitidis serogroup Ps (MnPs) A, C, Y, and W-135 were evaluated and it was determined that 50 ng/well of MnPs was optimal for the single-plex assay
Summary
Infections caused by the bacterium Neisseria meningitisdis remain an important medical need, in the developing world where periodic outbreaks are responsible for thousands of deaths [1]. Efficacious vaccines targeting four of these serogroups (A, C, Y and W135) have been developed by direct conjugation of polysaccharide antigens to protein carriers [3,4,5,6,7]. These vaccines provide benefit over traditional, unconjugated polysaccharide (Ps) vaccines by inducing T-cell memory, and afford long-term immunity in both children and adults [2]. Total serum-antibody specific for each of the Ps components of the vaccine under evaluation are usually assessed by enzyme-linked immunosorbent assay (ELISA). To create a higher throughput assay, we have developed an electrochemiluminescent (ECL)-based multi-plex assay for the quantitation of antibody responses to meningococcal polysaccharide from serogroups A, C, Y, and W-135
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