title: Quantitative measurements of N-linked glycoproteins in human plasma by SWATH-MS.
abstract: SWATH-MS is a data-independent acquisition method that generates, in a single measurement, a complete recording of the fragment ion spectra of all the analytes in a biological sample for which the precursor ions are within a predetermined m/z versus retention time window. To assess the performance and suitability of SWATH-MS-based protein quantification for clinical use, we compared SWATH-MS and SRM-MS-based quantification of N-linked glycoproteins in human plasma, a commonly used sample for biomarker discovery. Using dilution series of isotopically labeled heavy peptides representing biomarker candidates, the LOQ of SWATH-MS was determined to reach 0.0456 fmol at peptide level by targeted data analysis, which corresponds to a concentration of 5-10 ng protein/mL in plasma, while SRM reached a peptide LOQ of 0.0152 fmol. Moreover, the quantification of endogenous glycoproteins using SWATH-MS showed a high degree of reproducibility, with the mean CV of 14.90%, correlating well with SRM results (R(2) = 0.9784). Overall, SWATH-MS measurements showed a slightly lower sensitivity and a comparable reproducibility to state-of-the-art SRM measurements for targeted quantification of the N-glycosites in human blood. However, a significantly larger number of peptides can be quantified per analysis. We suggest that SWATH-MS analysis combined with N-glycoproteome enrichment in plasma samples is a promising integrative proteomic approach for biomarker discovery and verification.
authors: Department of Biology, Institute of Molecular Systems Biology, Zurich, Switzerland.
authors: Yansheng
authors: Y
authors: Liu
authors: Ruth
authors: R
authors: Hüttenhain
authors: Silvia
authors: S
authors: Surinova
authors: Ludovic C J
authors: LC
authors: Gillet
authors: Jeppe
authors: J
authors: Mouritsen
authors: Roland
authors: R
authors: Brunner
authors: Pedro
authors: P
authors: Navarro
authors: Ruedi
authors: R
authors: Aebersold
mesh_terms: D000595:Amino Acid Sequence; D015415:Biomarkers / Q000097:blood; D005260:Female; D006023:Glycoproteins / Q000097:blood* / Q000737:chemistry; D006031:Glycosylation; D006801:Humans; D007477:Ions; D008297:Male; D013058:Mass Spectrometry / Q000379:methods* / Q000592:standards; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D040901:Proteomics / Q000379:methods; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D015415:Biomarkers; D006023:Glycoproteins; D007477:Ions; D010455:Peptides