Microheterogeneity of a purified IgG1 due to asymmetric Fab glycosylation.
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.
Grebenau
R C
RC
Immunomedics Inc., Newark, NJ 07103.
Goldenberg
D M
DM
Chang
C H
CH
Koch
G A
GA
Gold
D V
DV
Kunz
A
A
Hansen
H J
HJ
D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry; D015153:Blotting, Western; D004591:Electrophoresis, Polyacrylamide Gel; D006031:Glycosylation; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry*; D007074:Immunoglobulin G / Q000737:chemistry*; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry; D066298:In Vitro Techniques; D046911:Macromolecular Substances; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase / Q000494:pharmacology; D051379:Mice
D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains; D046911:Macromolecular Substances; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase