iSwathX 2.0 for Processing DDA Spectral Libraries for DIA Data Analysis

Abstract

The iSwathX web application processes and normalizes mass spectrometry−based proteomics spectral libraries generated in the data‐dependent acquisition (DDA) approach. These libraries are stored in various proteomics repositories such as PeptideAtlas and NIST, or are user‐generated and provide reference data for data‐independent acquisition (DIA) targeted data extraction and analysis. iSwathX 2.0 can efficiently normalize DDA data from different instruments, gathered at different instances, and make it compatible with specific DIA experiments. Novel functions for parallel processing of DDA libraries and DIA report files, along with various data visualizations, are available in iSwathX 2.0. The step‐by‐step protocols provided here describe how the libraries are uploaded, processed, visualized, and downloaded using various modules of the application. They also provide detailed guidelines on the use of DIA report files for data analysis and visualization. © 2020 Wiley Periodicals LLC. Basic Protocol 1: Processing, combining, and visualizing two DDA libraries Basic Protocol 2: Parallel processing and combination of multiple DDA libraries Basic Protocol 3: Downstream processing, comparison, and visualization of DIA report files

PIONEER: Pipeline for Generating High‐Quality Spectral Libraries for DIA‐MS Data

“…For large-scale studies, libraries generated on different mass spectrometers under different LC conditions, and from different biological samples, tend to have differences in their retention times. Such libraries can be merged by iSwathX (Noor, Mohamedali, & Ranganathan, 2020), which creates a single unified library with the retention time alignment. Over the past years, several software tools have been developed for the generation of spectral libraries, such as SpectraST (Lam et al, 2007), X!Hunter (Craig, Cortens, Fenyo, & Beavis, 2006), Bibliospec (Frewen, Merrihew, Wu, Noble, & MacCoss, 2006), Pepitome (Dasari et al, 2012), and MSPepSearch (https: // chemdata.nist.gov/ ).…”

Advances in data‐independent acquisition mass spectrometry towards comprehensive digital proteome landscape

“…Several auxiliary tools have been reported for curation or statistical analysis of DIA data, including TRIC (Rost et al, 2016), iSwathX (Noor et al, 2020), Avant‐Grade (Vaca Jacome et al, 2020), DIALib‐QC, (Midha et al, 2020a) and others. Several tools have been developed to assist data analysis from the SWATH datasets‐one of the most popular DIA methods.…”

“…It was integrated into OpenSWATH and enables cross‐run alignment and peak‐picking from fragment ion data for quantification (Rost et al, 2016). iSwathX offers a web interface to combine spectral libraries and is compatible with the most commonly used DIA data analysis softwares such as Skyline, OpenSWATH, Spectronaut, and PeakView (Noor et al, 2020). The DIALib‐QC (DIA library quality control) is also integrated into SWATHAtlas (http://www.swathatlas.org) for systematic evaluation of a library's characteristics, completeness, and correctness across several parameters of compliance to ensure high‐quality library usage (Midha et al, 2020a).…”