title: Multiplexed analytical glycomics: rapid and confident IgG N-glycan structural elucidation.
abstract: N-glycans attached to the C(H)2 domains of the Fc or the antigen binding regions of IgG play an important role in stabilizing and modulating antibody activity. Exhaustive elucidation of 32 IgG N-glycans using a combination of weak anion exchange enrichment and exoglycosidase array digestion with subsequent profiling exceeded 48 h. Pursuing increased throughput and associated structural annotation confidence, we compared the 1.7 μm hydrophilic interaction phase for UPLC with CE-LIF for the rapid and comprehensive characterization of N-glycans released from healthy human serum polyclonal IgG. Combination of the data individually generated using each technique demonstrated that complete structural annotation was possible within a total analysis time of 20 min due to the advantageous orthogonality of the separation mechanisms. The parallel use of both analytical techniques provides a powerful platform for rapid and comprehensive analysis of IgG N-glycosylation present on therapeutic antibodies or on antibodies of biomedical or pathological significance.
authors.affiliation: NIBRT Dublin-Oxford Glycobiology Laboratory, NIBRT - The National Institute for Bioprocessing Research and Training, UCD Conway Institute of Biomolecular and Biomedical Research, University College Dublin, Belfield, Dublin 4, Ireland.
authors.forename: Stefan
authors.initials: S
authors.lastname: Mittermayr
authors.forename: Jonathan
authors.initials: J
authors.lastname: Bones
authors.forename: Margaret
authors.initials: M
authors.lastname: Doherty
authors.forename: András
authors.initials: A
authors.lastname: Guttman
authors.forename: Pauline M
authors.initials: PM
authors.lastname: Rudd
mesh_terms: D000328:Adult; D002853:Chromatography, Liquid; D019075:Electrophoresis, Capillary; D005260:Female; D054794:Glycomics; D006031:Glycosylation; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry*; D008297:Male; D011134:Polysaccharides / Q000737:chemistry*; D013050:Spectrometry, Fluorescence
chemical_list: D007074:Immunoglobulin G; D011134:Polysaccharides