Mass measurement accuracy of the Orbitrap in intact proteome analysis

• DOI: 10.1002/rcm.7574
• Publication Date: 2016-05-22T12:10:21Z
• Authors: Fan Yu; Houqin Fang; Kaijie Xiao; Yan Liu; Bingbing Xue; Zhixin Tian
• Keywords: Orbitrap; Proteome; Mass

Rationale The mass measurement accuracy (MMA) of Orbitrap spectrometers is 1–5 ppm according to the manufacturer's specification; yet, up 50 has been used as tolerance interpret data in literature. A systematic evaluation MMA thus necessary find optimal be used. Methods Reversed-phase liquid chromatography/tandem spectrometry (RPLC/MS/MS) analyses intact E. coli proteome were carried out on a Q Exactive spectrometer coupled Dionex UltiMate 3000 RSLCnano system. analysis included three technical replicates each day and was repeated for six continuous days right after calibration. obtained raw datasets searched using ProteinGoggle 2.0 under four different tolerances 5, 10, 15, 20 ppm. Results With both forward random database searches FDR ≤1% at spectrum level, most protein matches IDs 15 average precursor product ions from representative high, medium, low abundance proteins well common identified all 18 replicate runs found 0–4 ppm; no significant drift measured observed within calibration period 1 week. Conclusions Despite stated by manufacturer, identification ions. Weekly appropriate because 6-day period. Copyright © 2016 John Wiley & Sons, Ltd.