Determination of the epitope specificity of monoclonal antibodies against the inner core region of bacterial lipopolysaccharides by use of 3-deoxy-D-manno-octulosonate-containing synthetic antigens.
Partial structures of enterobacterial lipopolysaccharides (LPS) of the Rechemotype, consisting of lipid A and 3-deoxy-D-manno-2-octulosonic acid (Kdo), as well as oligosaccharides and derivative of Kdo were synthesized and used to characterize the epitope specificity of monoclonal antibodies against Re-mutant LPS. High-molecular-weight antigens, obtained after copolymerization of the respective allyl glycosides with acrylamide, and the haptenic oligosaccharides were used in immunoprecipitation, immune hemolysis, and in inhibition assays. A monoclonal antibody (clone 20, igM) recognizing a terminal Kdop group was shown to require for its binding the alpha-anomeric configuration and OH-4 and OH-5 groups, whereas the C-7 - C-8 chain was of minor importance. Another monoclonal antibody (clone 25, IgG3), which recognizes a (2--4)-linked Kdo disaccharide, was shown to require for its binding the alpha-anomeric configuration of both residues. The isomer having a reducing beta-Kdo residue was significantly less active, and that with a terminal beta-Kdo group was completely inactive. The OH-5 group of the reducing residue was shown to be not important for the specificity of this antibody, since it could be replaced by a hydrogen atom without loss of serological reactivity. The alpha-(2--8)-linked Kdo disaccharide was strongly cross-reactive with its (2--4)-linked isomer. The antibody recognized also parts of the 2-amino-2-deoxy-D-glucose backbone of lipid A.
Rozalski
A
A
Forschungsinstitut Borstel, Institut für Experimentelle Biologie und Medizin, F.R.G.
Brade
L
L
Kuhn
H M
HM
Brade
H
H
Kosma
P
P
Appelmelk
B J
BJ
Kusumoto
S
S
Paulsen
H
H
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000276:immunology*; D000918:Antibody Specificity; D002240:Carbohydrate Sequence; D000939:Epitopes / Q000276:immunology; D004926:Escherichia coli / Q000276:immunology; D006462:Hemolytic Plaque Technique; D066298:In Vitro Techniques; D008070:Lipopolysaccharides / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D011233:Precipitin Tests; D012475:Salmonella / Q000276:immunology; D012756:Sheep; D013400:Sugar Acids / Q000276:immunology*; D014614:Vaccines, Synthetic / Q000276:immunology
D000911:Antibodies, Monoclonal; D000939:Epitopes; D008070:Lipopolysaccharides; D013400:Sugar Acids; D014614:Vaccines, Synthetic; C002532:2-keto-3-deoxyoctonate