Glycosylation of Therapeutic Proteins: A Critical Quality Attribute -- Characterization of Protein Glycoforms at Intact Level by Orbitrap Mass Spectrometry -- Analysis of Intact Glycoproteins by Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry -- Fc Glycosylation Characterization of Human Immunoglobulins G using Immunocapture and LC-MS -- Fast Afucosylation Profiling of Glyco-Engineered Antibody Subunits by Middle Up Mass Spectrometry -- Characterization of Glycosylated Proteins at Subunit Level by HILIC/MS -- Analysis of Monoclonal Antibody Glycopeptides by Capillary Electrophoresis - Mass Spectrometry Coupling (CE-MS) -- Enrichment of Intact Glycopeptides using Strong Anion Exchange and Electrostatic Repulsion Hydrophilic Interaction Chromatography -- In-Depth Glycan Characterization of Therapeutic Glycoproteins by Stepwise PGC -SPE and LC-MS/MS -- Site-Specific N-glycosylation Analysis of Recombinant Proteins by LC/MSE -- Mapping O-glycosylation Sites using OpeRATOR and LC-MS -- Site-Specific O-Glycosylation Analysis by Liquid Chromatography/Mass Spectrometry with Electron-Transfer/Higher-Energy Collisional Dissociation -- Profiling of N-Linked Oligosaccharides of a Glycoprotein by UPLC-FLR-ESI-MS after Derivatization with Fluorescent Anthranilamide -- Evaluating N-glycosylation of a Therapeutic Monoclonal Antibody using UHPLC-FLR-MS with RapiFluor-MS Labeling -- Routine Analysis of N-glycans using Liquid Chromatography Coupled to Routine Mass Detection -- Online 2D-LC for Complex N-Glycan Analysis from Biopharmaceuticals -- Profiling, Relative Quantification, and Identification of Sialylated N-Linked Oligosaccharides by UPLC-FLR-ESI/MS after Derivatization with Fluorescent Anthranilamide -- Linkage Analysis of Oligo- and Polysaccharides: A Tutorial -- Use of Exoglycosidases for the Structural Characterization of Glycans -- Determination of Isomeric Glycan Structures by Permethylation and Liquid Chromatography-Mass Spectrometry (LC-MS) -- Optimization of Multiple Glycosidase and Chemical Stabilization Strategies for N-glycan Isomer Detection by Mass Spectrometry Imaging in Formalin-Fixed, Paraffin-Embedded Tissues -- Glycosylation Profiling of Glycoproteins Secreted from Cultured Cells Using Glycan Node Analysis and GC-MS -- Array-Based N-Glycan Profiling of Cells in Culture -- Analysis of Oligomeric and Glycosylated Proteins by Size-Exclusion Chromatography Coupled with Multi-Angle Light Scattering -- Analysis of Glycoproteins by ATR-FTIR Spectroscopy: Comparative Assessment -- Deploying Mass Spectrometric Data Analysis in the Amazon AWS Cloud Computing Environment.

This volume presents methods used for the analysis of glycoproteins at different levels-intact, subunit, glycopeptide, and monosaccharide--, and discusses and solves most analytical challenges that a scientist working on glycoproteins may come across. The chapters in this book cover topics such as the role of glycosylation on the properties of therapeutic glycoproteins; different analytical methods to characterize glycosylation, from the intact proteins to the glycan level, for both N-linked and O-linked glycoproteins; mass spectrometry imaging methodology for glycosylation analysis in tissues; approaches to characterizing glycosylation on cultured cells; and the use of cloud computing to deploy mass spectrometry data analysis. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Cutting-edge and thorough, Mass Spectrometry of Glycoproteins: Methods and Protocols is a valuable resource for scientists interested in learning more about this developing field.