Showing results for 
Search instead for 
Did you mean: 

Top-down analysis of monoclonal antibody IgG1 by electron transfer dissociation Orbitrap FTMS

Reputable Mentor II
Reputable Mentor II
Fornelli L, Damoc E, Thomas PM, Kelleher NL, Aizikov K, Denisov E, Makarov A, Tsybin YO.
Mol Cell Proteomics. 2012 Dec;11(12):1758-67.
Primary structure information of proteins employed as bio-therapeutics is essential to understand their structure-function relationship, in the rational design of new therapeutics, and for quality control. Given both the large size (around 150 kDa) and structural complexity of intact immunoglobulins G (IgGs), which include a variable number of disulfide bridges, their extensive fragmentation and subsequent sequence determination by tandem mass spectrometry (MS) are challenging. Here, we applied electron transfer dissociation (ETD) implemented on a hybrid Orbitrap Fourier transform mass spectrometer (FTMS) to analyze a commercial recombinant IgG in a liquid chromatography (LC)-tandem mass spectrometry (MS/MS) top-down experiment. The lack of sensitivity typically observed during top-down MS of large proteins was addressed by averaging time-domain transients recorded in different LC-MS/MS experiments before performing FT signal processing. The results demonstrate that improved signal-to-noise ratio combined with higher resolution and mass accuracy provided by Orbitrap FTMS (compared to previous applications of top-down ETD-based proteomics on IgGs) are essential for their comprehensive analysis. Specifically, ETD on Orbitrap FTMS produced about 33% sequence coverage of an intact IgG, signifying an almost two-fold increase in IgG sequence coverage in comparison with previous ETD-based analysis of intact monoclonal antibodies of a similar subclass. These results suggest the potential application of the developed methodology to other classes of large proteins and biomolecules.
Ecole Polytechnique Federale de Lausanne, Switzerland;
Version history
Last update:
‎10-15-2021 12:06 PM
Updated by: