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Comprehensive Characterization of Intact Monoclonal Antibody Using High Resolution Benchtop Quadrupole-Orbitrap LC-MS/MS

Reputable Mentor II
Reputable Mentor II
Hao Z, Zhang Y, Horn D, Jiang X, Wu SL, Huhmer A, Bennett PK.
Scientific Poster
Purpose: A LC/MS-based workflow solution was developed for robust, accurate and comprehensive intact monoclonal antibody (mAb) characterization. Methods: Thermo Scientific Q Exactive quadrupole-Orbitrap mass spectrometers were used for intact mass measurement and top-down sequencing. Full MS spectra of intact or reduced mAb were analyzed using Thermo Scientific Protein Deconvolution 2.0 software that utilizes the ReSpectTM algorithm for molecular mass determination. The top-down msx HCD spectra were analyzed using Thermo Scientific ProSight PC 2.0. Results: A mass error of less than 10 ppm was routinely achieved for intact mAb mass measurement. Low mass modifications, such as oxidation, can be confidently identified on substructure level such as intact Fab, or light chain. Using an on-line high resolution topdown MSMS approach, over 30% of the fragmentation site was covered for intact light chain as well as for Fab heavy chain. Sequence coverage from top-down approach also confirmed disulfide linkage on partially reduced samples.

Thermo Fisher Scientific
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Last update:
‎10-15-2021 07:47 AM
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