Run-Specific Calibration Transition Lists

Overview

Run-specific transition lists can be generated from search results of pseudo-DDA data (e.g. from DIA-Umpire or diaTracer) derived from the same DIA raw files. This approach can be used to improve the calibration of retention time, mass-to-charge and ion mobility values for each individual run.

OpenSWATH supports generic calibration transition lists in tab- or comma-separated value (TSV/CSV) and sqlite-based (PQP) formats. OpenSwathAssayGenerator is part of OpenMS and is available for Windows, macOS and Linux.

Contact and Support

FragPipe is supported by a very active community via their github issues tracker.

EasyPQP is supported by a very active community via their github issues tracker.

For OpenSwathAssayGenerator, we provide support via the OpenMS support channels.

Tutorial

The easiest way to generate run-specific transition lists is to use the FragPipe platform. The FragPipe team provides extensive documentation and tutorials on their website. Please follow their instructions to generate spectral libraries. The resulting *_run_peaks.tsv files will be in a tab delimited format that can be directly processed with the OpenSwathAssayGenerator tool in OpenMS ( follow the remaining steps in the Generic Transition Lists section).

OpenSwathWorkflow allows for an optional additional nonlinear calibration step (performed after linear alignment). You can reduce the *_run_peaks.tsv to a smaller set of peptides to be used for the linear calibration step by using EasyPQP.

run1_diau_run_peaks_file=sample_1_run_peaks.tsv
OpenSwathAssayGenerator -in $run1_diau_run_peaks_file -out sample_1_run_peaks_irt_nonlinear.pqp
easypqp reduce --in sample_1_run_peaks_irt_nonlinear.pqp --out sample_1_run_peaks_irt_linear.pqp --bins 10 --peptides 5