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The Impact II, a Very High-Resolution Quadrupole Time-of-Flight Instrument (QTOF) for Deep Shotgun Proteomics.

Beck S, Michalski A, Raether O, Lubeck M, Kaspar S, Goedecke N, Baessmann C, Hornburg D, Meier F, Paron I, Kulak NA, Cox J, Mann M - Mol. Cell Proteomics (2015)

Bottom Line: The new reflectron and detector improve resolving power compared with the previous model up to 80%, i.e. to 40,000 at m/z 1222.We analyzed the ion current from the inlet capillary and found very high transmission (>80%) up to the collision cell.Finally, after high pH reversed-phase fractionation we identified 9515 proteins in a triplicate measurement of HeLa peptide mixture and 11,257 proteins in single measurements of cerebellum-the highest proteome coverage reported with a QTOF instrument so far.

View Article: PubMed Central - PubMed

Affiliation: From the ‡Proteomics and Signal Transduction, Max-Planck-Institute of Biochemistry, Am Klopferspitz 18, 82152 Martinsried, Germany;

No MeSH data available.


Related in: MedlinePlus

Resolution and mass accuracy of A, a peptide isotope cluster (m/z 558.8063, r = 33k) and B, Fd unit of Adalimumab (m/z 25442.5157, r = 63k, 0.26 ppm). Overall improvement of the resolution with the improved detector, C, and the achieved mass accuracy dependent on the summed peptide intensity, D, in a shotgun proteomics experiment using the QTOF optimized version of MaxQuant.
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Figure 4: Resolution and mass accuracy of A, a peptide isotope cluster (m/z 558.8063, r = 33k) and B, Fd unit of Adalimumab (m/z 25442.5157, r = 63k, 0.26 ppm). Overall improvement of the resolution with the improved detector, C, and the achieved mass accuracy dependent on the summed peptide intensity, D, in a shotgun proteomics experiment using the QTOF optimized version of MaxQuant.

Mentions: In summary, the resolving power of the TOF analyzer is expected to increase by about 70 to 80% by the introduction of the new collision cell, reflectron and detector. To test this experimentally on a standard proteomic sample, we analyzed data from a HeLa digest. Resolution for typical peptides is in excess of 33,000 as illustrated by an example in Fig. 4A. Further increase in the resolution can be obtained by the “Focus mode,” which involves real time processing and alignment of successive pulses and increases accuracy of flight time determination, when multiple ions of one species reach the detector at the same time (45). This can be helpful to resolve the isotope distributions of proteins, as shown in Fig. 4B, which features a resolution in excess of 60,000 for an antibody subunit (> 25 kDa). In TOF measurements, resolution tends to increase with m/z (Fig. 4C), and reached more than 40,000 for the TuneMix component at m/z 1222. This constitutes a 70 to 80% improvement over the previous impact model, the impact HD.


The Impact II, a Very High-Resolution Quadrupole Time-of-Flight Instrument (QTOF) for Deep Shotgun Proteomics.

Beck S, Michalski A, Raether O, Lubeck M, Kaspar S, Goedecke N, Baessmann C, Hornburg D, Meier F, Paron I, Kulak NA, Cox J, Mann M - Mol. Cell Proteomics (2015)

Resolution and mass accuracy of A, a peptide isotope cluster (m/z 558.8063, r = 33k) and B, Fd unit of Adalimumab (m/z 25442.5157, r = 63k, 0.26 ppm). Overall improvement of the resolution with the improved detector, C, and the achieved mass accuracy dependent on the summed peptide intensity, D, in a shotgun proteomics experiment using the QTOF optimized version of MaxQuant.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4587313&req=5

Figure 4: Resolution and mass accuracy of A, a peptide isotope cluster (m/z 558.8063, r = 33k) and B, Fd unit of Adalimumab (m/z 25442.5157, r = 63k, 0.26 ppm). Overall improvement of the resolution with the improved detector, C, and the achieved mass accuracy dependent on the summed peptide intensity, D, in a shotgun proteomics experiment using the QTOF optimized version of MaxQuant.
Mentions: In summary, the resolving power of the TOF analyzer is expected to increase by about 70 to 80% by the introduction of the new collision cell, reflectron and detector. To test this experimentally on a standard proteomic sample, we analyzed data from a HeLa digest. Resolution for typical peptides is in excess of 33,000 as illustrated by an example in Fig. 4A. Further increase in the resolution can be obtained by the “Focus mode,” which involves real time processing and alignment of successive pulses and increases accuracy of flight time determination, when multiple ions of one species reach the detector at the same time (45). This can be helpful to resolve the isotope distributions of proteins, as shown in Fig. 4B, which features a resolution in excess of 60,000 for an antibody subunit (> 25 kDa). In TOF measurements, resolution tends to increase with m/z (Fig. 4C), and reached more than 40,000 for the TuneMix component at m/z 1222. This constitutes a 70 to 80% improvement over the previous impact model, the impact HD.

Bottom Line: The new reflectron and detector improve resolving power compared with the previous model up to 80%, i.e. to 40,000 at m/z 1222.We analyzed the ion current from the inlet capillary and found very high transmission (>80%) up to the collision cell.Finally, after high pH reversed-phase fractionation we identified 9515 proteins in a triplicate measurement of HeLa peptide mixture and 11,257 proteins in single measurements of cerebellum-the highest proteome coverage reported with a QTOF instrument so far.

View Article: PubMed Central - PubMed

Affiliation: From the ‡Proteomics and Signal Transduction, Max-Planck-Institute of Biochemistry, Am Klopferspitz 18, 82152 Martinsried, Germany;

No MeSH data available.


Related in: MedlinePlus