This study examines the effects of precursor and fragment ion mass accuracy on protein identification using the Accurate Mass Q-TOF. Results demonstrate that fragment ion mass accuracy is as important (if not more important) than precursor ion mass accuracy in reducing false positive identification rates in MS/MS database searches, and plays a critical role in improving the throughput of proteomics research projects.
This document is an application note from Agilent Technologies that investigates the effect of fragment ion mass accuracy on protein identification rates in shotgun proteomics. It compares the performance of the Agilent 6510 Q-TOF LC/MS system against an Agilent 6330 Ion Trap mass spectrometer using HeLa cell samples.
The study demonstrates that high fragment ion mass accuracy is as important as precursor ion mass accuracy in reducing false positive identification rates. The authors utilized the 6510 Q-TOF with specific mass tolerances (10 ppm for precursors and 40 ppm for fragments) to achieve higher identification yields and confidence compared to the ion trap system.
| Acquisition rate | 20 spectra/sec |
|---|---|
| Resolving power | 15,000 |
| Dynamic range | 3.5 orders of magnitude |
| Precursor ion tolerance (Q-TOF) | 10 ppm |
| Fragment ion tolerance (Q-TOF) | 40 ppm |
| Precursor ion tolerance (Ion Trap) | 2.5 Da |
| Fragment ion tolerance (Ion Trap) | 0.7 Da |
| MS Scan range (Q-TOF) | 300-2000 m/z |
| MS/MS Scan range (Q-TOF) | 50-3000 m/z |
| MS Scan range (Ion Trap) | 300-1800 m/z |
| MS/MS Scan range (Ion Trap) | 100-2000 m/z |
The optimal precursor ion tolerance for the 6510 Q-TOF is 10 ppm.
The optimal fragment ion tolerance for the 6510 Q-TOF is 40 ppm.
Yes, high fragment ion mass accuracy is as important as precursor ion mass accuracy in reducing false positive identification rates.