Carolina Silva1,2, Cátia Santa1,3, Sandra Isabel Anjo1,4, Bruno Manadas1. 1. CNC - Center for Neuroscience and Cell Biology, University of Coimbra, Coimbra, Portugal. 2. Department of Chemistry, University of Aveiro, Aveiro, Portugal. 3. Institute for Interdisciplinary Research, University of Coimbra, Coimbra, Portugal. 4. Faculty of Sciences and Technology, University of Coimbra, Coimbra, Portugal.
Abstract
PURPOSE: Peripheral blood mononuclear cells (PBMCs) play quite diverse and important roles in monitoring immune homeostasis. Thus, this subset of blood cells may provide access to potential physiological relevant biomolecules, namely proteins. For this reason, PBMCs represent a promising alternative biological sample in scientific research, particularly as a source of potential biomarkers. Prior proteomic studies of PBMCs from healthy individuals focused only on a qualitative analysis, lacking the quantitative analysis information crucial for biomarker discovery, since most of the biological alterations result in slight changes in protein levels, not affecting the overall proteome composition. Therefore, this study aimed to provide a comprehensive PBMCs proteome library to be use in protein quantification by SWATH-MS. EXPERIMENTAL DESIGN: A SWATH-MS library was generated by a comprehensive 2D-LC-MS/MC analysis of a pooled sample of PBMCs from 6 different donors. GeLC-SWATH-MS analysis of the 6 donors was further used to test the generated library. RESULTS: The generated library comprises 1102 proteins involved in diverse human diseases and in immune system-related pathways. When tested in biological samples this library allowed the quantification of 920 different proteins, and around 700 per individual. CONCLUSIONS AND CLINICAL RELEVANCE: The provided PBMCs proteome library will be useful in further studies that aim to reproducibly quantify a large number of PBMCs' proteins without the need to perform protein identification. Furthermore, this robust microLC-SWATH-MS analysis is suitable with clinical practice.
PURPOSE: Peripheral blood mononuclear cells (PBMCs) play quite diverse and important roles in monitoring immune homeostasis. Thus, this subset of blood cells may provide access to potential physiological relevant biomolecules, namely proteins. For this reason, PBMCs represent a promising alternative biological sample in scientific research, particularly as a source of potential biomarkers. Prior proteomic studies of PBMCs from healthy individuals focused only on a qualitative analysis, lacking the quantitative analysis information crucial for biomarker discovery, since most of the biological alterations result in slight changes in protein levels, not affecting the overall proteome composition. Therefore, this study aimed to provide a comprehensive PBMCs proteome library to be use in protein quantification by SWATH-MS. EXPERIMENTAL DESIGN: A SWATH-MS library was generated by a comprehensive 2D-LC-MS/MC analysis of a pooled sample of PBMCs from 6 different donors. GeLC-SWATH-MS analysis of the 6 donors was further used to test the generated library. RESULTS: The generated library comprises 1102 proteins involved in diverse human diseases and in immune system-related pathways. When tested in biological samples this library allowed the quantification of 920 different proteins, and around 700 per individual. CONCLUSIONS AND CLINICAL RELEVANCE: The provided PBMCs proteome library will be useful in further studies that aim to reproducibly quantify a large number of PBMCs' proteins without the need to perform protein identification. Furthermore, this robust microLC-SWATH-MS analysis is suitable with clinical practice.
Authors: Johannes Nemeth; Valentina Vongrad; Karin J Metzner; Victoria P Strouvelle; Rainer Weber; Patrick Pedrioli; Ruedi Aebersold; Huldrych F Günthard; Ben C Collins Journal: Mol Cell Proteomics Date: 2017-02-21 Impact factor: 5.911