A repository of assays to quantify 10,000 human proteins by SWATH-MS

Abstract

Mass spectrometry is the method of choice for deep and reliable exploration of the (human) proteome. Targeted mass spectrometry reliably detects and quantifies pre-determined sets of proteins in a complex biological matrix and is used in studies that rely on the quantitatively accurate and reproducible measurement of proteins across multiple samples. It requires the one-time, a priori generation of a specific measurement assay for each targeted protein. SWATH-MS is a mass spectrometric method that combines data-independent acquisition (DIA) and targeted data analysis and vastly extends the throughput of proteins that can be targeted in a sample compared to selected reaction monitoring (SRM). Here we present a compendium of highly specific assays covering more than 10,000 human proteins and enabling their targeted analysis in SWATH-MS datasets acquired from research or clinical specimens. This resource supports the confident detection and quantification of 50.9% of all human proteins annotated by UniProtKB/Swiss-Prot and is therefore expected to find wide application in basic and clinical research. Data are available via ProteomeXchange (PXD000953-954) and SWATHAtlas (SAL00016-35).

Analysis of Major Histocompatibility Complex (MHC) Immunopeptidomes Using Mass Spectrometry*

“…To extract quantitative information from such digital SWATH-MS data, high-quality assay libraries are required (109,111). Such libraries contain retention-time and fragmentation information of the peptides to be quantified.…”

Spectral Libraries for SWATH‐MS Assays for Drosophila melanogaster and Solanum lycopersicum

“…When changing the q-value, we observed an increase in the number of peptides identified with a correlated increase of the coefficient of variation ( Figure 2A and 2B and Figure S3). However, even with a FDR of 5%, we found the technical reproducibility to be similar (median CVs below 8.5% for Drosophila and tomato) to other studies [9,24].Next, we assessed the performance of SWATH-MS analysed with our spectral libraries compared to other DDA mode on the same or different MS platforms. For this comparison, a Drosophila embryo sample was injected four times on a Q Exactive (Fisher Scientific, Waltham, MA, USA), a LTQ-Orbitrap Velos (Fisher Scientific, Waltham, MA, USA) or a TripleTOF 6600 mass spectrometer, all in DDA acquisition mode, and on a TripleTOF 6600 in SWATH acquisition mode (Supporting Information).…”

“…The spectral libraries and all massspectrometry data are available in the MassIVE repository with the dataset identifiers MSV000081074 and MSV000081075 and the PRIDE repository with the dataset identifiers PXD006493 and PXD006495. For DIA, the spectral library contains experimental acquired spectra of the precursors and can be produced directly from the SWATH data, by reconstituting the MSMS spectra using the retention time of the precursors and the fragments [7], or by DDA analysis of the sample, which can be fractionated to increase the number of spectra in the library [9]. Ideally, the spectral library should be generated on a MS instrument similar to the one used to acquire further SWATH-MS data as the correlation of the fragment intensities for a peptide acquired on different instrument was shown to be rather low [10].…”

“…Ideally, the spectral library should be generated on a MS instrument similar to the one used to acquire further SWATH-MS data as the correlation of the fragment intensities for a peptide acquired on different instrument was shown to be rather low [10]. Although Rosenberger et al recently published a library containing assays for 10,000 human proteins [9] and deep libraries for other applications such as SRM (Selected Reaction Monitoring) have been generated [11][12][13], the number of publicly available spectral libraries for SWATH-MS is still limited. Several other studies have produced libraries for other species [14][15][16].…”

The Obama Administration's Cancer Moonshot: A Call for Proteomics

“…Advancements in technology and informatics now enable us to quantify in excess of 10,000 proteins in a single experiment, half of all human proteins annotated by the UniProt knowledge base (5). We now have CLIA/CAP-accredited proteomic assays today that can quantitatively measure the phosphorylation levels of most of the "actionable" drug targets, from small biopsies, often requiring less material than required for next-generation sequencing/genomic analysis.…”