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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., et al. (2015). The Impact II, a Very High-Resolution Quadrupole Time-of-Flight Instrument (QTOF) for Deep Shotgun Proteomics. MOLECULAR & CELLULAR PROTEOMICS, 14(7), 2014-2029. doi:10.1074/mcp.M114.047407.

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 Creators:
Beck, Scarlet1, Author           
Michalski, Annette2, Author
Raether, Oliver2, Author
Lubeck, Markus2, Author
Kaspar, Stephanie2, Author
Goedecke, Niels2, Author
Baessmann, Carsten2, Author
Hornburg, Daniel1, Author           
Meier, Florian1, Author           
Paron, Igor1, Author           
Kulak, Nils A.1, Author           
Cox, Juergen3, Author           
Mann, Matthias1, Author           
Affiliations:
1Mann, Matthias / Proteomics and Signal Transduction, Max Planck Institute of Biochemistry, Max Planck Society, ou_1565159              
2external, ou_persistent22              
3Cox, Jürgen / Computational Systems Biochemistry, Max Planck Institute of Biochemistry, Max Planck Society, ou_2063284              

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Free keywords: SPECTROMETRY-BASED PROTEOMICS; TANDEM MASS-SPECTROMETRY; BOTTOM-UP PROTEOMICS; ELECTROSPRAY-IONIZATION; LARGE BIOMOLECULES; QUANTIFICATION; PERFORMANCE; PEPTIDES; PROTEIN; CELLS
 Abstract: Hybrid quadrupole time-of-flight (QTOF) mass spectrometry is one of the two major principles used in proteomics. Although based on simple fundamentals, it has over the last decades greatly evolved in terms of achievable resolution, mass accuracy, and dynamic range. The Bruker impact platform of QTOF instruments takes advantage of these developments and here we develop and evaluate the impact II for shotgun proteomics applications. Adaption of our heated liquid chromatography system achieved very narrow peptide elution peaks. The impact II is equipped with a new collision cell with both axial and radial ion ejection, more than doubling ion extraction at high tandem MS frequencies. 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. Simulation and measurement indicated 60% transfer into the flight tube. We adapted MaxQuant for QTOF data, improving absolute average mass deviations to better than 1.45 ppm. More than 4800 proteins can be identified in a single run of HeLa digest in a 90 min gradient. The workflow achieved high technical reproducibility (R2 > 0.99) and accurate fold change determination in spike-in experiments in complex mixtures. Using label-free quantification we rapidly quantified haploid against diploid yeast and characterized overall proteome differences in mouse cell lines originating from different tissues. 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.

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Language(s): eng - English
 Dates: 2015
 Publication Status: Issued
 Pages: 16
 Publishing info: -
 Table of Contents: -
 Rev. Type: Peer
 Identifiers: ISI: 000357434400021
DOI: 10.1074/mcp.M114.047407
 Degree: -

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Title: MOLECULAR & CELLULAR PROTEOMICS
Source Genre: Journal
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Publ. Info: 9650 ROCKVILLE PIKE, BETHESDA, MD 20814-3996 USA : AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
Pages: - Volume / Issue: 14 (7) Sequence Number: - Start / End Page: 2014 - 2029 Identifier: ISSN: 1535-9476