A synthetic glycoconjugate representing the genus-specific epitope of chlamydial lipopolysaccharide exhibits the same specificity as its natural counterpart.
The tetrasaccharide 3-deoxy-alpha-D-manno-2-octulosonic acid (alpha-KDO) (2----8)-alpha-KDO(2----4)-alpha-KDO(2----6)-beta GlcNAc, a partial structure of chlamydial lipopolysaccharide (LPS) representing a genus-specific epitope, was synthesized and covalently linked to bovine serum albumin, resulting in an artificial glycoconjugate antigen. Mice were immunized with the glycoconjugate to prepare chlamydia-specific monoclonal antibodies. They were selected with chlamydia-specific LPS antigens and the structurally and antigenically related Re-type LPS of a Salmonella minnesota rough mutant. Characterization of the selected antibodies was by (i) hemagglutination of sheep erythrocytes coated with recombinant chlamydia-specific LPS, (ii) inhibition by synthetic polyacrylamide derivatives containing the genus-specific epitope or partial structures thereof, (iii) enzyme immunoassay with recombinant LPS and synthetic bovine serum albumin glycoconjugates as solid-phase antigens, (iv) immunofluorescence of L929 monolayers infected with Chlamydia psittaci or C. trachomatis, and (v) Western immunoblots with glycoconjugates and LPS as the antigen. Two groups of monoclonal antibodies were obtained; the monoclonal antibodies in one group cross-reacted with chlamydial and Re-type LPS, but those of the other group were chlamydia specific. Among the latter, KDO trisaccharide-specific antibodies that had the same epitope specificity as antibodies obtained after immunization with chlamydial elementary bodies were identified; however, they exhibited a more than 100-fold higher affinity. In addition, antibodies that bound preferentially to the 2.8-linked KDO disaccharide were detected, although with lower affinity. The data show that the artificial glycoconjugate antigen is similar to its natural counterpart.
Fu
Y
Y
Division of Biochemical Microbiology, Forschungsinstitut Borstel, Germany.
Baumann
M
M
Kosma
P
P
Brade
L
L
Brade
H
H
D000818:Animals; D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial / Q000276:immunology; D015153:Blotting, Western; D002240:Carbohydrate Sequence; D002689:Chlamydia / Q000276:immunology*; D000939:Epitopes / Q000276:immunology*; D005455:Fluorescent Antibody Technique; D006001:Glycoconjugates / Q000138:chemical synthesis / Q000276:immunology*; D006386:Hemagglutination Tests; D007124:Immunoenzyme Techniques; D008070:Lipopolysaccharides / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D013400:Sugar Acids / Q000276:immunology
D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D000939:Epitopes; D006001:Glycoconjugates; D008070:Lipopolysaccharides; D013400:Sugar Acids; C002532:2-keto-3-deoxyoctonate