Microheterogeneity of a purified IgG1 due to asymmetric Fab glycosylation

• Published in: Molecular immunology Vol. 29; no. 6; p. 751
• Main Authors: Grebenau, R C, Goldenberg, D M, Chang, C H, Koch, G A, Gold, D V, Kunz, A, Hansen, H J
• Format: Journal Article
• Language: English
• Published: England 01.06.1992
• Subjects: Animals; Antibodies, Monoclonal - chemistry; Blotting, Western; Electrophoresis, Polyacrylamide Gel; Glycosylation; Immunoglobulin Fab Fragments - chemistry; Immunoglobulin G - chemistry; Immunoglobulin Heavy Chains - chemistry; In Vitro Techniques; Macromolecular Substances; Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase - pharmacology; Mice
• ISSN: 0161-5890
• DOI: 10.1016/0161-5890(92)90185-Z

A murine monoclonal anti-granulocyte IgG1, IMMU-MN3, was seen to exhibit heterogeneity. On reduced SDS-PAGE, the purified antibody appeared as two heavy-chain bands of unequal intensity, and only one light-chain band. Hydrophobic interaction chromatography (HIC) also resolved two populations of the IMMU-MN3 antibody. Based on Concanavalin A affinity chromatography, enzymatic digestion with Endoglycosidase F and carbohydrate analysis, it was found that the heterogeneity detected by SDS-PAGE and HIC was due to differences in glycosylation. Furthermore, sequential gel analysis (non-reduced/reduced) demonstrated that the upper heavy-chain band was asymmetrically glycosylated.