Previous studies with group C meningococcal polysaccharide-tetanus toxoid (GCMP-TT) conjugates had suggested that the GCMP O-acetyl group masked the protective epitope for group C meningococci through steric hindrance or altered conformations. For this report, we confirmed this phenomenon and performed comparative studies with group Y meningococcal polysaccharide (GYMP)-TT to determine whether it might extend to other serogroups. The de-O-acetylated (dOA) polysaccharides (PSs) resulted in higher serum bactericidal activities (SBA) towards the O-acetylated (OA) meningococcal strains from the respective serogroups. High-resolution H-nuclear magnetic resonance spectroscopy at 500 MHz and competitive inhibition serum bactericidal assays were used to characterize the nature of the protective epitope. In head-to-head comparisons with OA PSs as SBA inhibitors, the dOA PSs provided 10 to 1,000 times better inhibition for GCMP in human and mouse antisera and 6 to 13 times better inhibition for GYMP in mouse antisera, using OA strains in all assays. In addition, the SBA for OA strains was highly correlated with dOA PS-specific immunoglobulin G (r=0.72 to 0.98) for both GCMP and GYMP. The results suggest that there may be a generalized role for the O-acetyl group to provide an epitope of misdirected immunogenicity for meningococcal PS capsules, enabling escape from immune surveillance. In addition to greater chemical consistency, the dOA forms of GCMP and GYMP conjugate vaccines endow greater immunologic competence to the PSs, rendering them capable of eliciting higher levels of functional antibodies toward the protective epitopes.