Definition of Brucella A and M epitopes by monoclonal typing reagents and synthetic oligosaccharides.
The paradigm that Brucella A and M epitopes are simultaneously expressed on single cells and within one antigen molecule was reinvestigated by using polysaccharide-specific murine monoclonal antibodies. Monoclonal antibodies were generated to the M antigen of Brucella melitensis 16M. Chemically defined lipopolysaccharides and O polysaccharides from Brucella abortus 1119-3, B. melitensis 16M, and Yersinia enterocolitica O:9 were used to dissect the binding profiles of the B. melitensis antibodies and an additional set of antibodies available from a B. abortus fusion experiment. Binding specificities were rationalized in terms of prototype A- and M-antigen structures, an interpretation supported by competitive binding studies with O polysaccharides and synthetic oligosaccharide analogs of the A and M antigens. Three binding patterns were characterized. Antibodies specific for the A antigen required five contiguous alpha 1,2-linked 4,6-dideoxy-4-formamido-D-mannopyranosyl residues, while antibodies with equal affinities for A or M epitopes were effectively inhibited by alpha 1,2-linked tri- or tetrasaccharides. Specificity for the M epitope correlated with binding of a critical disaccharide element alpha-D-Rha4NFo(1----3)alpha-D-Rha4NFo bracketed by alpha 1,2-linked residues. The binding profiles of Brucella monoclonal antibodies were consistent with the concept of simultaneous expression of A and M epitopes within a single molecule. A epitopes were present in the M antigen, and the discovery of isolated alpha 1,3 linkages in the A antigen suggests that M epitopes occur in all A antigens. Three monoclonal antibodies are proposed as standard reagents for the detection and identification of Brucella A and M antigens.
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D R
DR
Division of Biological Sciences, National Research Council of Canada, Ottawa, Ontario.
Cherwonogrodzky
J W
JW
Gidney
M A
MA
Meikle
P J
PJ
Perry
M B
MB
Peters
T
T
D000818:Animals; D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D000942:Antigens, Bacterial / Q000032:analysis* / Q000276:immunology; D001666:Binding Sites, Antibody; D001667:Binding, Competitive; D002002:Brucella / Q000276:immunology*; D002003:Brucella abortus / Q000276:immunology; D000939:Epitopes / Q000032:analysis* / Q000276:immunology; D005260:Female; D005779:Immunodiffusion; D007124:Immunoenzyme Techniques; D007202:Indicators and Reagents; D008070:Lipopolysaccharides / Q000032:analysis / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D015345:Oligonucleotide Probes; D012703:Serotyping / Q000379:methods
D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D000939:Epitopes; D007202:Indicators and Reagents; D008070:Lipopolysaccharides; D015345:Oligonucleotide Probes