Analysis of tandem mass spectra by FTMS for improved large-scale proteomics with superior protein quantification.
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 (approximately 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 (approximately 30% improvement for CAD and approximately 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.
McAlister
Graeme C
GC
Department of Chemistry, University of Wisconsin, Madison, Wisconsin 53706, USA.
Phanstiel
Doug
D
Wenger
Craig D
CD
Lee
M Violet
MV
Coon
Joshua J
JJ
D005583:Fourier Analysis; D011506:Proteins / Q000737:chemistry*; D040901:Proteomics / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*
D011506:Proteins