Quantitative Proteomics on Immune Cells of ME/CFS Patients Using SWATH-MS, 2025, Kumar, Peppercorn, Kleffmann

Quantitative Proteomics on Immune Cells of ME/CFS Patients Using SWATH-MS
Kumar, Abhishek; Peppercorn, Katie; Kleffmann, Torsten

Proteomics is one of the “omics” disciplines that has provided molecular insights into the pathophysiology of myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS).

Here we describe a complete SWATH-MS workflow for the quantitative profiling of proteins extracted from peripheral mononuclear blood cells to investigate proteomic alterations in ME/CFS. This workflow covers all steps of sample preparation, data acquisition, and data analysis. We describe the process of generating a comprehensive spectral library from a pre-fractionated peptide reference sample followed by the acquisition of DIA data sets of individual samples using a 5600+ TripleTOF mass spectrometer.

Examples of both library-based and library-free data analysis pipelines are presented based on the PeakView/MarkerView software package (commercial) and DIA-NN (free) software respectively.

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Katie Peppercorn is from the Department of Biochemistry, University of Otago, Dunedin, New Zealand and has coauthored papers with Warren Tate (and edited the Springer Protocols book on ME/CFS research techniques with Warren).

The other two researchers are from the Centre for Protein Research, Research Infrastructure Centre, University of Otago, Dunedin, New Zealand.