Preparation and characterization of monoclonal antibodies to an N-linked oligosaccharide.
Two monoclonal antibodies to an N-linked oligosaccharide, MT-5 and MT-9, have been prepared by immunization with a pyridylaminated, asialylated, galactosylated, fucosylated, bisected biantennary sugar. The reactivity of these antibodies was monitored by their reaction with human asialoglycophorin in a solid-phase enzyme-linked immunosorbent assay. Both antibodies reacted with the sugar chains of various human glycoproteins such as immunoglobulin G, transferrin, gamma-glutamyl transpeptidase, alpha 1-acid glycoprotein, and alpha-fetoprotein. Treatment of asialoglycophorin with beta-N-acetylhexosaminidase or alpha-mannosidase resulted in reduction of the binding to these antibodies. The reactivity of MT-5 to asialoglycophorin was slightly inhibited by D-mannose and N-acetylglucosamine, whereas that of MT-9 was inhibited by D-mannose, N-acetyl-D-glucosamine, chitobiose, and L-fucose. The epitope specificity of MT-5 appears to be a sugar chain containing biantennary N-acetyl-D-glucosamine residues, the bisected N-acetyl-D-glucosamine residue, and a trimannosyl core. The epitope to which MT-9 is directed may be a complex made up of beta-mannose, chitobiose, and L-fucose. These studies indicate that immunization with pyridylaminated sugars can produce antibodies that recognize N-linked oligosaccharides. Monoclonal/polyclonal antibodies to the N-linked sugar chains of glycopeptides would be useful in such studies of proteins.
Masutani
S
S
Second Department of Surgery, Osaka University Medical School, Japan.
Miyazawa
N
N
Fujii
S
S
Nishikawa
A
A
Matsukawa
H
H
Shimano
T
T
Mori
T
T
Taniguchi
N
N
D000911:Antibodies, Monoclonal / Q000032:analysis / Q000302:isolation & purification*; D000918:Antibody Specificity; D000937:Antigen-Antibody Reactions; D001212:Asialoglycoproteins / Q000276:immunology; D001710:Biotin; D015153:Blotting, Western; D002241:Carbohydrates / Q000276:immunology; D005959:Glucosidases / Q000276:immunology; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D006868:Hydrolysis; D007074:Immunoglobulin G / Q000276:immunology; D007202:Indicators and Reagents; D009844:Oligosaccharides / Q000276:immunology*
D000911:Antibodies, Monoclonal; D001212:Asialoglycoproteins; D002241:Carbohydrates; D007074:Immunoglobulin G; D007202:Indicators and Reagents; D009844:Oligosaccharides; C026503:asialoglycophorin; D001710:Biotin; D005959:Glucosidases