Abstract
Higher–order antibody aggregates remain challenging to characterize using conventional mass spectrometry because of their low abundance, broad mass distributions, and extensive heterogeneity. Here, the practical performance of a commercial electrostatic linear ion trap charge detection mass spectrometer (XevoTM CDMS) was evaluated for therapeutic antibody analysis. The platform provided reproducible mass and charge measurements, with repeatability and intermediate-precision RSDs below 1%, and maintained consistent mass determination across the concentration and solution conditions examined. Controlled mixtures of antibody-derived species showed a linear molar-ratio-dependent ion response, supporting semi–quantitative estimation of relative species abundance under defined conditions. For heat-stressed trastuzumab, CDMS measurements were qualitatively consistent with SEC–MALS, mass photometry, and conventional native MS, while enabling direct mass determination of individual ions within highly heterogeneous high–molecular–weight (HMW) populations that exhibited extensive charge–state overlap and could not be reliably deconvoluted by conventional native MS. High–duty–cycle acquisition increased sampling of low–abundance, high–m/z ions and improved access to higher-order aggregate signals. Joint single ion mass–charge analysis further differentiated statistical ion families within the broad HMW fraction and related major mass differences to independently obtained compositional information. These results support commercial XevoTM CDMS as a practical multidimensional platform for antibody aggregate characterization.