Articles by Alexander Greenshields-Watson in JoVE
Using X-ray Crystallography, Biophysics, and Functional Assays to Determine the Mechanisms Governing T-cell Receptor Recognition of Cancer Antigens Bruce J. MacLachlan*1, Alexander Greenshields-Watson*1, Georgina H Mason*1, Andrea J Schauenburg1, Valentina Bianchi1,2,3, Pierre J Rizkallah1, Andrew K Sewell1, Anna Fuller1, David K Cole1 1Division of Infection and Immunity and Systems Immunity Research Institute, Cardiff University, 2Department of Oncology, University Hospital of Lausanne (CHUV), 3Ludwig Insitutue for Cancer Research, Lausanne Branch, University of Lausanne Here, we describe methods that we commonly employ in the laboratory to determine how the nature of the interaction between the T-cell receptor and tumor antigens, presented by human leukocyte antigens, governs T-cell functionality; these methods include protein production, X-ray crystallography, biophysics, and functional T-cell experiments.
Other articles by Alexander Greenshields-Watson on PubMed
T-cell Libraries Allow Simple Parallel Generation of Multiple Peptide-specific Human T-cell Clones Journal of Immunological Methods. Mar, 2016 | Pubmed ID: 26826277 Isolation of peptide-specific T-cell clones is highly desirable for determining the role of T-cells in human disease, as well as for the development of therapies and diagnostics. However, generation of monoclonal T-cells with the required specificity is challenging and time-consuming. Here we describe a library-based strategy for the simple parallel detection and isolation of multiple peptide-specific human T-cell clones from CD8(+) or CD4(+) polyclonal T-cell populations. T-cells were first amplified by CD3/CD28 microbeads in a 96U-well library format, prior to screening for desired peptide recognition. T-cells from peptide-reactive wells were then subjected to cytokine-mediated enrichment followed by single-cell cloning, with the entire process from sample to validated clone taking as little as 6 weeks. Overall, T-cell libraries represent an efficient and relatively rapid tool for the generation of peptide-specific T-cell clones, with applications shown here in infectious disease (Epstein-Barr virus, influenza A, and Ebola virus), autoimmunity (type 1 diabetes) and cancer.