Affinity chromatography for purification of antibodies to Neisseria gonorrhoeae and Neisseria meningitidis lipopolysaccharides.

Abstract

Lipopolysaccharides (LPSs) were prepared by phenol-water extraction of the gonococcal strain 8551 and the group B meningococcal strain 44/76, digested with pronase, and purified by ultracentrifugation and Sepharose CL-6B fractionation in the presence of 1.5 per cent sodium deoxycholate. On SDS-PAGE with 10 per cent acrylamide the purified 125I-labelled LPSs migrated as single, low-molecular-weight components. The LPSs were coupled to CNBr-activated Sepharose 4B for affinity purification of antibodies to the common antigenic factor 1 and the sero-type factor 5 of LPS 8551, and antibodies to LPS 44/76. The antibodies eluted showed ELISA activity against wells coated with LPS or whole cells of the bacteria, the antibody activity being inhibited by LPS. SDS-PAGE of whole cells of the strain 8551 and immunoblotting with the anti-factor 1 or -factor 5 antibodies resulted in single, broad bands corresponding to the low-molecular-weight LPS subunits.