A 30 kD protein was isolated from an outer membrane vesicle (OMV) preparation of Neisseria meningitidis using single-step preparative electrophoresis. From a total of 23.4 mg of OMV-applied on SDS-PAGE, 3.9 mg of 30 kD protein were recovered in the single step of fractionation. This corresponds to 17% of the total protein applied. The protein was homogeneous by SDS-PAGE and N-terminal (ASELGsRSPYYVQAD) amino acid sequence analysis. Monoclonal antibodies to the 30 kD protein were raised in mice, confirming its purity and specificity Sera from N. meningitidis patients reacted with the 30 kD protein, but this did not occur with normal healthy individuals. Thus the method used allowed the purification of the antigen in a high yield, suggesting that it can be used as an alternative approach for the purification of other very hydrophobic proteins.

Two monoclonal antibodies (mAbs) were produced against a serogroup B Neisseria meningitidis strain. These mAbs recognized two epitopes in the class 5 outer membrane proteins (OMP), designated P5.7 and P5.Bm, and were able to kill the homologous strain through complement activation. Both epitopes were surface exposed and 68% of group B meningococcal clinical isolates had one or both epitopes present in their class 5 OMP. Antibodies to one or both epitopes were demonstrated in 17 patients with meningococcal meningitis using an ELISA inhibition assay. Of the 17 paired sera, 41% and 29% of the acute-phase sera had antibodies to the P5.7 and P5.Bm epitopes, respectively. Immunoglobulin G to P5.Bm were found in all 17 convalescent-phase sera while specific antibodies against P5.7 were only found in 6 of these sera. These results demonstrate the potential importance of the P5.Bm and P5.7 epitopes on the class 5 OMP as candidates for vaccine composition.