Analysis of Tandem Mass Spectra by FTMS for Improved Large-Scale Proteomics with Superior Protein Quantification

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Abstract
Using a newly developed dual-cell quadrupole linear ion trap−orbitrap hybrid mass spectrometer (dcQLT−orbitrap), we demonstrate the utility of collecting high-resolution tandem mass spectral data for large-scale shotgun proteomics. Multiple nanoLC−MS/MS experiments on both an older generation quadrupole linear ion trap−orbitrap hybrid (QLT−orbitrap) and the dcQLT−orbitrap, using both resonant-excitation CAD and beam-type CAD (HCD), were performed. Resulting from various technological advances (e.g., a stacked ring ion guide AP inlet, a dual cell QLT), the dcQLT−orbitrap exhibited increased duty cycle (∼1.5−2 times) and sensitivity for both CAD (ion trap detection) and HCD (orbitrap detection) methods. As compared to the older system, the dcQLT−orbitrap produced significantly more unique peptide identifications for both methods (∼30% improvement for CAD and ∼115% improvement for HCD). The sizable improvement of the HCD method on the dcQLT−orbitrap system outperforms the current standard method of CAD with ion trap detection for large-scale analysis. Finally, we demonstrate that the increased HCD performance translates to a direct and substantial improvement in protein quantitation precision using isobaric tags.