title: Screening method using selected reaction monitoring for targeted proteomics studies of nasal lavage fluid.
abstract: Proteomic-based studies of nasal lavage fluid (NLF) may identify molecular pathways associated with disease pathology and new biomarker candidates of upper airway diseases. However, most studies have used rather tedious untargeted MS techniques. Selected reaction monitoring (SRM) is a sensitive and specific technique that can be used with high throughput. In this study, we developed a semiquantitative SRM-based method targeting 244 NLF proteins. The protein set was identified through a literature study in combination with untargeted LC-MS/MS analyses of trypsin-digested NLF samples. The SRM assays were designed using MS/MS data either downloaded from a proteomic data repository or experimentally obtained. Each protein is represented by one to five peptides, resulting in 708 SRM assays. Three to four transitions per assay were used to ensure analyte specificity. The majority (69%) of the assays showed good within-day precision (coefficient of variation ≤ 20%). The accuracy of the method was evaluated by analyzing four samples prepared with varying amounts of four proteins. Peptide and protein ratios were in good agreement with expected ratios. In conclusion, a high throughput screening method for relative quantification of 244 NLF proteins was developed. The method should be of general use in any proteomic study of the upper airways.
authors.affiliation: Department of Laboratory Medicine, Lund, Division of Occupational and Environmental Medicine, Lund University, SE-221 85 Lund, Sweden. harriet.mortstedt@med.lu.se
authors.forename: Harriet
authors.initials: H
authors.lastname: Mörtstedt
authors.forename: Monica H
authors.initials: MH
authors.lastname: Kåredal
authors.forename: Bo A G
authors.initials: BA
authors.lastname: Jönsson
authors.forename: Christian H
authors.initials: CH
authors.lastname: Lindh
mesh_terms: D015415:Biomarkers; D002853:Chromatography, Liquid; D057166:High-Throughput Screening Assays; D006801:Humans; D013058:Mass Spectrometry; D017817:Nasal Lavage Fluid / Q000737:chemistry*; D010455:Peptides / Q000737:chemistry / Q000145:classification / Q000302:isolation & purification; D011506:Proteins / Q000737:chemistry / Q000145:classification / Q000302:isolation & purification; D040901:Proteomics
chemical_list: D015415:Biomarkers; D010455:Peptides; D011506:Proteins