Monoclonal antibodies against the heptose region of enterobacterial lipopolysaccharides

Abstract

Murine monoclonal antibodies (MAbs) against the rough mutant lipopolysaccharides (LPS) of Salmonella minnesota strain R4 and R7 (chemotypes Rd₂P ^- and Rd₁P^-, respectively) were obtained after immunization of BALB/c and NZB mice, respectively, with heat-killed bacteria. A total of 5 MAbs was obtained which were serologically characterized by hemagglutination, EIA and Western blot using LPS, de-O-acylated LPS and dephosphorylated LPS. In addition, the completely deacylated carbohydrate backbone, the deacylated, dephosphorylated and reduced carbohydrate backbone, and synthetic partial structures were covalently linked to bovine serum albumin resulting in artificial glycoconjugate antigens. Both groups of MAbs (anti-R4 and anti-R7) were highly specific for the Rd₂P^- and Rd₁P^- chemotypes, respectively, since they did not react with LPS of the Ra, Rb, Rc or Re chemotype. The R4 antibodies required for binding the core region [composed of 3-deoxy-α-D-manno-octulosonic acid (Kdo) and L-glycero-α-D- manno-heptopyranose (Hep)] and the phosphorylated glucosamine backbone of the lipid A moiety. The R7 antibodies bound to some extent to the core oligosaccharide Hep(1→3)Hep(1→5)Kdo(2→6), however, better binding was observed when the glucosamine backbone of lipid A was also present. In this case however, phosphoryl groups were not required. These antibodies can detect enterobacteria with an LPS of the Rd-chemotype, and may be useful to characterize also other naturally occurring rough mutants or genetically manipulated strains used to clone genes of the rfa locus.