| Reference |
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| Reference Type | Literature | IEDB_Reference:1038233 |
| Title | Mild Acid Elution and MHC Immunoaffinity Chromatography Reveal Similar Albeit Not Identical Profiles of the HLA Class I Immunopeptidome. | |
| Authors | Theo Sturm; Benedikt Sautter; Tobias P Wörner; Stefan Stevanović; Hans-Georg Rammensee; Oliver Planz; Albert J R Heck; Ruedi Aebersold | |
| Affiliations | Institute of Molecular Systems Biology, Department of Biology, ETH Zürich, 8093 Zürich, Switzerland; Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Utrecht University, 3584 CH Utrecht, The Netherlands; Netherlands Proteomics Centre, 3584 CH Utrecht, The Netherlands; Philochem AG, 8112 Otelfingen, Switzerland; Department of Immunology, Institute for Cell Biology, University of Tübingen, 72076 Tübingen, Germany; Faculty of Science, University of Zurich, 8057 Zürich, Switzerland. | |
| Journal | J Proteome Res | |
| Year | 2021 | |
| Abstract | To understand and treat immunology-related diseases, a comprehensive, unbiased characterization of major histocompatibility complex (MHC) peptide ligands is of key importance. Preceding the analysis by mass spectrometry, MHC class I peptide ligands are typically isolated by MHC immunoaffinity chromatography (MHC-IAC) and less often by mild acid elution (MAE). MAE may provide a cheap alternative to MHC-IAC for suspension cells but has been hampered by the high number of contaminating, MHC-unrelated peptides. Here, we optimized MAE, yielding MHC peptide ligand purities of more than 80%. When compared with MHC-IAC, obtained peptides were similar in numbers, identities, and to a large extent intensities, while the percentage of cysteinylated peptides was 5 times higher in MAE. The latter benefitted the discovery of MHC-allotype-specific, distinct cysteinylation frequencies at individual positions of MHC peptide ligands. MAE revealed many MHC ligands with unmodified, N-terminal cysteine residues which get lost in MHC-IAC workflows. The results support the idea that MAE might be particularly valuable for the high-confidence analysis of post-translational modifications by avoiding the exposure of the investigated peptides to enzymes and reactive molecules in the cell lysate. Our improved and carefully documented MAE workflow represents a high-quality, cost-effective alternative to MHC-IAC for suspension cells. | |
| Curation Last Updated | 2025-02-11 00:23:08 | |