Aman Makaju1; Kim Alving2; Bing Wang2; Rosa Viner1
ASMS 2020
Purpose: Correctly and confidently identify N-glycosylation sites and its glycoforms in a biologic using High-Field Asymmetric Waveform Ion Mobility Spectrometry System (FAIMS Pro) interfaced to the Orbitrap Eclipse Mass Spectrometer.
Methods: LC/MS analysis was performed on Thermo Scientific™ Orbitrap Eclipse™ platform coupled with Thermo Scientific™ FAIMS Pro™ interface. The raw files were analyzed using Thermo Scientific™ Proteome Discoverer™ 2.4 software with Byonicnode, and Byologic®, or Thermo Scientific™ BioPharmaFinder™ 4.0 software.
Results: We successfully identified 7 glycosylation sites including N383 in IgE.Using HCD or ETD fragmentation in combination with optimized FAIMS CV settings we were able to identify significantly more different glycoforms than without FAIMS. Overall, the FAIMS-LC-MS/MS workflow significantly improves glycoform profiling and characterization in heavily glycosylated proteins without enrichment or pre-fractionation.
1ThermoFisher Scientific, San Jose, CA; 2Sanofi, Waltham, MA
United States
