Mass spectrometry–based identification of MHC-bound peptides for immunopeptidomics
Peptide antigens bound to molecules encoded by the major histocompatibility complex (MHC) and presented on the cell surface form the targets of T lymphocytes. This critical arm of the adaptive immune system facilitates the eradication of pathogen-infected and cancerous cells, as well as the producti...
| Main Authors: | , , |
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| Format: | Journal article |
| Language: | English |
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Springer Nature
2019
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| _version_ | 1797101201635934208 |
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| author | Purcell, A Ramarathinam, S Ternette, N |
| author_facet | Purcell, A Ramarathinam, S Ternette, N |
| author_sort | Purcell, A |
| collection | OXFORD |
| description | Peptide antigens bound to molecules encoded by the major histocompatibility complex (MHC) and presented on the cell surface form the targets of T lymphocytes. This critical arm of the adaptive immune system facilitates the eradication of pathogen-infected and cancerous cells, as well as the production of antibodies. Methods to identify these peptide antigens are critical to the development of new vaccines, for which the goal is the generation of effective adaptive immune responses and long-lasting immune memory. Here, we describe a robust protocol for the identification of MHC-bound peptides from cell lines and tissues, using nano-ultra-performance liquid chromatography coupled to high-resolution mass spectrometry (nUPLC–MS/MS) and recent improvements in methods for isolation and characterization of these peptides. The protocol starts with the immunoaffinity capture of naturally processed MHC-peptide complexes. The peptides dissociate from the class I human leukocyte antigens (HLAs) upon acid denaturation. This peptide cargo is then extracted and separated into fractions by HPLC, and the peptides in these fractions are identified using nUPLC–MS/MS. With this protocol, several thousand peptides can be identified from a wide variety of cell types, including cancerous and infected cells and those from tissues, with a turnaround time of 2–3 d. |
| first_indexed | 2024-03-07T05:48:29Z |
| format | Journal article |
| id | oxford-uuid:e805f2e4-749a-4204-8238-c292c9179ece |
| institution | University of Oxford |
| language | English |
| last_indexed | 2024-03-07T05:48:29Z |
| publishDate | 2019 |
| publisher | Springer Nature |
| record_format | dspace |
| spelling | oxford-uuid:e805f2e4-749a-4204-8238-c292c9179ece2022-03-27T10:43:34ZMass spectrometry–based identification of MHC-bound peptides for immunopeptidomicsJournal articlehttp://purl.org/coar/resource_type/c_dcae04bcuuid:e805f2e4-749a-4204-8238-c292c9179eceEnglishSymplectic Elements at OxfordSpringer Nature2019Purcell, ARamarathinam, STernette, NPeptide antigens bound to molecules encoded by the major histocompatibility complex (MHC) and presented on the cell surface form the targets of T lymphocytes. This critical arm of the adaptive immune system facilitates the eradication of pathogen-infected and cancerous cells, as well as the production of antibodies. Methods to identify these peptide antigens are critical to the development of new vaccines, for which the goal is the generation of effective adaptive immune responses and long-lasting immune memory. Here, we describe a robust protocol for the identification of MHC-bound peptides from cell lines and tissues, using nano-ultra-performance liquid chromatography coupled to high-resolution mass spectrometry (nUPLC–MS/MS) and recent improvements in methods for isolation and characterization of these peptides. The protocol starts with the immunoaffinity capture of naturally processed MHC-peptide complexes. The peptides dissociate from the class I human leukocyte antigens (HLAs) upon acid denaturation. This peptide cargo is then extracted and separated into fractions by HPLC, and the peptides in these fractions are identified using nUPLC–MS/MS. With this protocol, several thousand peptides can be identified from a wide variety of cell types, including cancerous and infected cells and those from tissues, with a turnaround time of 2–3 d. |
| spellingShingle | Purcell, A Ramarathinam, S Ternette, N Mass spectrometry–based identification of MHC-bound peptides for immunopeptidomics |
| title | Mass spectrometry–based identification of MHC-bound peptides for immunopeptidomics |
| title_full | Mass spectrometry–based identification of MHC-bound peptides for immunopeptidomics |
| title_fullStr | Mass spectrometry–based identification of MHC-bound peptides for immunopeptidomics |
| title_full_unstemmed | Mass spectrometry–based identification of MHC-bound peptides for immunopeptidomics |
| title_short | Mass spectrometry–based identification of MHC-bound peptides for immunopeptidomics |
| title_sort | mass spectrometry based identification of mhc bound peptides for immunopeptidomics |
| work_keys_str_mv | AT purcella massspectrometrybasedidentificationofmhcboundpeptidesforimmunopeptidomics AT ramarathinams massspectrometrybasedidentificationofmhcboundpeptidesforimmunopeptidomics AT ternetten massspectrometrybasedidentificationofmhcboundpeptidesforimmunopeptidomics |