Book/Journal Article

DKFZ-2025-02133

http://join2-wiki.gsi.de/foswiki/pub/Main/Artwork/join2_logo100x88.png High-Coverage Immunopeptidomics Using timsTOF Mass Spectrometers with Thunder-DDA-PASEF Boosted by MS2Rescore.

Gómez-Zepeda, D. (First author)DKFZ* ; Beyrle, J.DKFZ* ; Preikschat, A. ; Declercq, A. ; Chen, Y.DKFZ* ; Gabriels, R. ; Martens, L. ; Distler, U. ; Tenzer, S. (Last author)DKFZ*

2026
Humana Press Totowa, NJ
ISBN: 978-1-0716-4831-5 (print), 978-1-0716-4832-2 (electronic)

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Please use a persistent id in citations: doi:10.1007/978-1-0716-4832-2_5  doi:DOI:10.1007/978-1-0716-4832-2_5

Abstract: Major histocompatibility complex (MHC, or human leukocyte antigen, HLA) peptide ligands can be exploited to develop immunotherapies targeting immunogenic disease-specific immunopeptides, such as virus- or cancer mutation-derived peptides. Liquid chromatography coupled with mass spectrometry (LC-MS)-based immunopeptidomics is the gold standard for identifying MHC ligands. We previously optimized a workflow enabling the identification of more than 10,000 MHC class I ligands per cell line. This process comprises three major steps: (I) a high-recovery immunopeptidome enrichment, (II) an optimized MS acquisition in the timsTOF Pro called Thunder-Data-Dependent Acquisition with Parallel Accumulation-SErial Fragmentation (Thunder-DDA-PASEF), and (III) peptide identification using PEAKS XPro boosted by MS2Rescore data-driven rescoring. Here, we describe our workflow for deep-coverage immunopeptidomics step-by-step, from sample preparation to data analysis and validation.

Keyword(s): Humans (MeSH) ; Peptides: immunology (MeSH) ; Peptides: chemistry (MeSH) ; Peptides: metabolism (MeSH) ; Chromatography, Liquid: methods (MeSH) ; Proteomics: methods (MeSH) ; Mass Spectrometry: methods (MeSH) ; Workflow (MeSH) ; Histocompatibility Antigens Class I: immunology (MeSH) ; Histocompatibility Antigens Class I: metabolism (MeSH) ; Ligands (MeSH) ; Tandem Mass Spectrometry: methods (MeSH) ; Human leucocyte antigen ; Immunopeptidomics ; Immunoprecipitation ; Major histocompatibility complex ; Mass spectrometry ; Re-scoring ; Sample preparation ; Peptides ; Histocompatibility Antigens Class I ; Ligands

Note: #EA:D190#LA:D191# / First Online: 15 October 2025

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Contributing Institute(s):

1. HI-TRON zentral (D190)
2. Hi-TRON Immunoproteomik (D191)

Research Program(s):

1. 314 - Immunologie und Krebs (POF4-314) (POF4-314)

Appears in the scientific report 2025

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Database coverage:
; SCOPUS

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