IgG Monoclonal Antibodies: Analytical Perspectives

Abstract

IgG1 and IgG4 monoclonal antibodies (mAbs) represent the two most widely used IgG subclasses in therapeutic applications. This review provides a comprehensive overview of analytical characterization tools essential for understanding the physicochemical and functional properties of these molecules. The importance of robust analytical techniques is highlighted for structural conformation, posttranslational modifications (PTMs), charge heterogeneity, and aggregation behaviour. Advanced methodologies including imaged capillary isoelectric focusing (icIEF), membrane-confined electrophoresis (MCE), and LC-MS/MS peptide mapping are discussed for their roles in providing high-resolution insights into charge variants and PTM hotspots. The section on LC-MS/MS explores the advantages of different fragmentation techniques and the utility of middle-up approaches in profile consistency across samples. Advances in multidimensional LC-MS/MS workflows demonstrate improved efficiency in critical quality attribute detection. Complementary techniques such as size-exclusion chromatography coupled with multi-angle light scattering (SEC-MALS) and dynamic light scattering (DLS) are employed to quantify colloidal stability and evaluate global conformational changes. Additionally, the significance of in silico modelling and molecular dynamics simulations for predicting degradation-prone regions and electrostatic compatibility are discussed. Overall, the review underscores the integration of analytical strategies and modelling tools crucial for the developability engineering and biosimilarity assessment of IgG1 and IgG4 monoclonal antibodies, ensuring product quality and safety in therapeutic applications for researchers and biopharmaceutical developers.