Epitope size, specificity and equilibrium constant for four monoclonal antibodies binding to the O:4 polysaccharide antigen of Salmonella serogroup B bacteria.
One rat (MAST 83) and three mouse (MAST 107, 108 and 112) monoclonal antibodies (mAbs) directed against Salmonella serogroup BO lipopolysaccharide (LPS) were characterized and found to bind to the O:4 epitope but recognizing different surfaces of the polysaccharide chain. The epitopes were defined from the combined results of: (i) binding specificities in enzyme immuno assay (EIA) against chemically defined LPS and glycoconjugates; (ii) studies of affinity constants in Farr-assay for binding to oligosaccharides purified from LPS, or chemically synthesized; and (iii) knowledge of the conformation of the O-polysaccharide chain of Salmonella BO bacteria. Two of the antibodies, MAST 83 and 108, bound to the O:4 epitope when present in the terminal non-reducing end as well as an intrachain determinant of the O-polysaccharide, whereas MAST 107 and 112 bound only to the O:4 epitope when present as an intrachain determinant. The equilibrium constants (K values), determined for binding of the mAbs and a Fab-fragment isolated from one of them to a 125l-labelled tyramine-derivative of a Salmonella BO dodecasaccharide, were: 4.3 x 10(5) (MAST 83), 1.0 x 10(5) (MAST 107), 1.3 x 10(5) (MAST 107 Fab), 4.5 x 10(4) (MAST 108) and 1.9 x 10(5) l/mol (MAST 112). The results suggest that each epitope encompasses the O:4 specifying D-abequosyl residue together with different numbers of saccharides varying in size from di- to tetrasaccharides from the linear backbone chain. The antibodies also bind to different surfaces of the O-polysaccharide chain as suggested by its conformation.
Lind
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Department of Clinical Bacteriology, Karolinska Institute, Huddinge Hospital, Sweden.
Lindberg
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D000907:Antibodies, Bacterial / Q000276:immunology*; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000915:Antibody Affinity; D000918:Antibody Specificity; D002240:Carbohydrate Sequence; D000939:Epitopes; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000737:chemistry / Q000276:immunology; D011135:Polysaccharides, Bacterial / Q000276:immunology*; D012475:Salmonella / Q000276:immunology*; D012486:Salmonella typhimurium / Q000276:immunology
D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D000939:Epitopes; D009844:Oligosaccharides; D011135:Polysaccharides, Bacterial