Use of single chain MHC technology to investigate co-agonism in human CD8+ T cell activation
| Autor: | Paul A. MacAry, Keith G. Gould, Joanna Ai Ling Choo, Ee Chee Ren, Maryam Hamidinia, Nicholas R. J. Gascoigne, Zi Zong Ho, Antonio Bertoletti, Joanna Brzostek, Chien Tei Too, Xiang Zhao |
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| Jazyk: | angličtina |
| Rok vydání: | 2019 |
| Předmět: |
0301 basic medicine
Agonist medicine.drug_class T cell General Chemical Engineering chemical and pharmacologic phenomena CD8-Positive T-Lymphocytes Major histocompatibility complex Lymphocyte Activation Transfection General Biochemistry Genetics and Molecular Biology 03 medical and health sciences 0302 clinical medicine MHC class I medicine Cytotoxic T cell Humans biology General Immunology and Microbiology Chemistry General Neuroscience Histocompatibility Antigens Class I Cell biology 030104 developmental biology medicine.anatomical_structure Cell culture 030220 oncology & carcinogenesis biology.protein CD8 |
| Popis: | Non-stimulatory self peptide MHC (pMHC) complexes do not induce T cell activation and effector functions, but can enhance T cell responses to agonist pMHC, through a process termed co-agonism. This protocol describes an experimental system to investigate co-agonism during human CD8+ T cell activation by expressing human MHC class I molecules presenting pre-determined peptides as single polypeptides (single chain MHC) in a xenogeneic cell line. We expressed single chain MHCs under conditions where low levels of agonist single chain p-MHC complexes and high levels of non-stimulatory single chain p-MHC complexes were expressed. Use of this experimental system allowed us to compare CD8+ T cell responses to agonist pMHC in the presence or absence of non-stimulatory pMHC. The protocol describes cell line transfection with single chain MHC constructs, generation of stable cell lines, culture of hepatitis B virus-specific human CD8+ T cells and T cell activation experiments simultaneously quantifying cytokine production and degranulation. The presented methods can be used for research on different aspects of CD8+ T cell activation in human T cell systems with known peptide MHC specificity. |
| Databáze: | OpenAIRE |
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