New Application Note on Antibody Oxidation Analysis

A Rapid LC-MS Assay for Monitoring of mAb Oxidation at the Subunit Level
Methionine oxidation is considered a critical quality attribute of therapeutic antibodies and may impact the clinical safety and efficacy. Therefore, monitoring of methionine oxidations is required during the discovery, development, and production of therapeutic antibodies. Traditional methods to characterize oxidation rely on tryptic peptide mapping and LC-MS, a labor intensive and time- consuming process that generates large data sets and requires trained and skilled manual interpretations.

In this application note, we present an LC-MS assay based on sample preparation with specific enzymatic digestion using FabRICATOR and deglycosylation using GlycINATOR that enables rapid antibody oxidation analysis. This middle-level oxidation assay is applied to a wide range of antibodies. Details of the optimized mobile phase, chromatography gradients, and mass spectrometry settings are given. The middle-level workflow can be used to study many critical quality attributes and the assay can be considered a platform method for therapeutic antibody characterization.
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FabRICATOR (IdeS) – Below hinge digestion of IgG.
GlycINATOR (EndoS2) – Hydrolysis of all Fc-glycans.