title: Glycosylation profiling of a therapeutic recombinant monoclonal antibody with two N-linked glycosylation sites using liquid chromatography coupled to a hybrid quadrupole time-of-flight mass spectrometer.
abstract: Monoclonal antibodies have been used increasingly as therapeutic agents to target various diseases. Although most monoclonal antibodies have only one N-linked glycosylation site in the Fc region, N-linked glycosylation sites in the Fab region have also been observed. Because glycosylation of a monoclonal antibody can have a significant impact on its effector function, efficacy, clearance, and immunogenicity, it is essential to assess the glycosylation profile during cell line and clone selection studies and to assess the impact of cell culture conditions on the glycoform distribution during process optimization studies to ensure that the antibody is being produced with appropriate and consistent glycosylation. This article describes a liquid chromatography-mass spectrometry-based approach, in combination with papain digestion and partial reduction, to obtain site-specific glycosylation profile information for a therapeutic monoclonal antibody with two N-linked glycosylation sites in the heavy chain.
authors.affiliation: Biopharmaceutical Research and Development, Eli Lilly and Company, Indianapolis, IN 46285, USA.
authors.forename: Amareth
authors.initials: A
authors.lastname: Lim
authors.forename: Angelia
authors.initials: A
authors.lastname: Reed-Bogan
authors.forename: Bryan J
authors.initials: BJ
authors.lastname: Harmon
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000378:metabolism* / Q000627:therapeutic use; D001665:Binding Sites; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D002853:Chromatography, Liquid; D006031:Glycosylation; D007140:Immunoglobulin Fab Fragments / Q000378:metabolism; D007141:Immunoglobulin Fc Fragments / Q000378:metabolism; D007143:Immunoglobulin Heavy Chains / Q000378:metabolism; D007147:Immunoglobulin Light Chains / Q000378:metabolism; D008239:Lysine / Q000378:metabolism; D010084:Oxidation-Reduction; D010206:Papain / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry* / Q000378:metabolism* / Q000627:therapeutic use; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization
chemical_list: D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D007141:Immunoglobulin Fc Fragments; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D011994:Recombinant Proteins; D010206:Papain; D008239:Lysine