MONOCLONAL-ANTIBODIES AGAINST HAEMOPHILUS LIPOPOLYSACCHARIDES - CLONEDP8 SPECIFIC FOR HAEMOPHILUS-DUCREYI AND CLONE DH24 BINDING TO LACTO-N-NEOTETRAOSE

Mouse monoclonal antibodies (MAbs) DP8 [immunoglobulin G1(kappa)] and DH24 [immunoglobulin M(kappa)], which are specific for Haemophilus duc reyi lipopolysaccharide (LPS), were generated by fusing mouse myeloma NS0 cells with spleen cells of BALB/c mice immunized with a total memb rane preparation of H. ducreyi. MAb DP8 reacted in whole-cell enzyme i mmunoassay (EIA) and colony dot immunoblotting with all 50 strains of H. ducreyi but not with any other bacteria tested, which suggests an e xposed and species-specific epitope on the H. ducreyi cell surface. Th is conclusion was supported by the finding that DP8 bound to all six H . ducreyi LPSs tested but not to any of the Haemophilus influenzae or enterobacterial LPSs or synthetic glycoconjugates. The MAb DH24 bound to 43 of 50 strains of H. ducreyi and to few strains of H. influenzae, Neisseria gonorrhoeae, and Neisseria meningitidis, as evaluated by wh ole cell EIA and colony dot immunoblotting. The MAb DH24 reacted with five of the six H. ducreyi LPSs tested and with the lacto-N-neotetraos e (Gal beta 1-->4GlcNAc beta 1-->3Gal beta 1-->4Glc) series of synthet ic glycoconjugates, as determined by EIA. By using polysaccharides obt ained after both mild acidic hydrolysis and strong alkali treatment an d dephosphorylated samples as inhibitors of the MAbs binding to H. duc reyi LPS antigens, it could be shown that phosphate groups were essent ial for the binding of DP8 to LPS but that they did not affect antigen ic recognition by DH24. None of the MAbs bound to isolated lipid A, bu t aggregation caused by the fatty acids of lipid A was essential for e pitope recognition.