Mass spectrometry for the biophysical characterization of therapeutic monoclonal antibodies

Monoclonal antibodies (mAbs) are powerful therapeutics, and their characterization has drawn considerable attention and urgency. Unlike small‐molecule drugs (150–600 Da) that have rigid structures, mAbs (∼150 kDa) are engineered proteins that undergo complicated folding and can exist in a number of low‐energy structures, posing a challenge for traditional methods in structural biology. Mass spectrometry (MS)‐based biophysical characterization approaches can provide structural information, bringing high sensitivity, fast turnaround, and small sample consumption. This review outlines various MS‐based strategies for protein biophysical characterization and then reviews how these strategies provide structural information of mAbs at the protein level (intact or top‐down approaches), peptide, and residue level (bottom‐up approaches), affording information on higher order structure, aggregation, and the nature of antibody complexes.