Protein Therapeutics: Antibody Characterization with ECD

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Labile post-translational modifications are better preserved using ECD than conventional MS fragmentation techniques such as collision-induced dissociation (CID). Common modifications, such as glycosylation and phosphorylation, can impact the safety, efficacy, and binding activity of monoclonal antibodies but are challenging to identify. Stephen Sciuto from Agilent Technologies and colleagues analyzed tryptic digests of NIST mAb and Infliximab using an 6545XT Q-TOF mass spectrometer in ExD or CID mode. ECD provided higher sequence coverage than CID, reaching 100% for glycosylated peptides in Infliximab. The ability to pinpoint glycosylation sites using fragment ions demonstrates the power of ECD for characterizing glycopeptides.