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Reputable Mentor II
Reputable Mentor II
Gallien S, Duriez E, Crone C, Kellmann M, Moehring T, Domon B.
Mol Cell Proteomics. 2012 Dec;11(12):1709-23.
There is an immediate need for improved methods to systematically and precisely quantify large sets of peptides in complex biological samples. While to date protein quantification in biological samples is routinely performed on triple quadrupole instruments operated in selected reaction monitoring mode (SRM), two major challenges remain. Firstly, the number of peptides to be included in one survey experiment needs to be increased to routinely reach several hundreds, and secondly, the degree of selectivity should be improved to reliably discriminate the targeted analytes from background interferences. High resolution and accurate mass (HR/AM) analysis on the recently developed Q‐Exactive mass spectrometer can potentially address these issues. This instrument presents a unique configuration: it is constituted of the orbitrap mass analyzer equipped with a quadrupole mass filter as front‐end for precursor ion mass selection. This configuration enables new quantitative methods based on HR/AM measurements, including targeted analysis in MS mode (single ion monitoring, SIM) and in MS/MS mode (parallel reaction monitoring, PRM). While the ability of the quadrupole to select a restricted m/z range allows overcoming the dynamic range limitations associated with trapping devices, the MS/MS mode provides an additional stage of selectivity. When applied to targeted protein quantification in urine samples and benchmarked with the reference SRM technique, the quadrupole orbitrap instrument exhibits similar or better performances in terms of selectivity, dynamic range, and sensitivity. These high performances are further enhanced by leveraging the multiplexing capability of the instrument to design novel acquisition methods and apply them to large targeted proteomic studies for the first time, as demonstrated on 770 tryptic yeast peptides analyzed in one 60 minute experiment. The increased quality of quadrupole‐orbitrap data has the potential to improve existing protein quantification methods in complex samples and to cope with the pressing demand of systems biology or biomarker evaluation studies.
Luxembourg Clinical Proteomics Center (LCP), CRP‐Santé, Strassen, Luxembourg
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