Analysis of glycosylation changes in IgG using lectins.
A simple rapid assay based on the ability of lectins to bind carbohydrates has been developed to analyse changes in the oligosaccharide chains of IgG. Bandeiraea simplicifolia lectin and Ricinus communis agglutinin have been used to detect terminal N-acetylglucosamine and galactose moieties respectively in IgG using immunodot-blotting. IgG samples (approximately 1 micrograms) were dot-blotted onto nitrocellulose followed by boiling of the blots to expose the carbohydrate moieties. The blots were then treated with biotinylated lectins followed by either streptavidin-biotin-hydrogen peroxidase conjugate or 125I-labelled streptavidin. The colour was developed using chloronaphthol and the blots read on a densitometer. The labelled blots were cut and read on a gamma counter. The use of a monoclonal antibody to N-acetylglucosamine is also discussed. The results obtained using this method are comparable to those obtained by structural analysis.
Department of Immunology, University College and Middlesex School of Medicine, London, U.K.
N
N
Sumar
K B
KB
Bodman
T W
TW
Rademacher
R A
RA
Dwek
P
P
Williams
R B
RB
Parekh
J
J
Edge
G A
GA
Rook
D A
DA
Isenberg
F C
FC
Hay
D001172:Arthritis, Rheumatoid / Q000276:immunology; D002240:Carbohydrate Sequence; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D006023:Glycoproteins / Q000032:analysis; D006031:Glycosylation; D006801:Humans; D007124:Immunoenzyme Techniques; D007074:Immunoglobulin G / Q000032:analysis* / Q000378:metabolism; D037102:Lectins / Q000032:analysis; D008969:Molecular Sequence Data; D037121:Plant Lectins; D012276:Ricin / Q000378:metabolism
D006023:Glycoproteins; C045475:Griffonia simplicifolia lectins; D007074:Immunoglobulin G; D037102:Lectins; D037121:Plant Lectins; D012276:Ricin