Dunbar, RVerdon, Daniel2016-12-062016http://hdl.handle.net/2292/31236This research project built on previous findings within the host laboratory that naïve CD4+ and CD8+ T cells can be stimulated and maintained in vitro under the influence of the common γc cytokine IL-7 and retain central memory characteristics. Further, this project aimed to expand these observations to the production of single-cell derived CD4+ and CD8+ T cell clones of known and unknown epitope specificity. We hypothesised that administration of cytokines IL-7 and IL-21 during T cell cloning would allow retention of ‘central memory’ characteristics and decouple proliferation from differentiation. The specific aims of this research project were to investigate the functional and phenotypic consequences of naïve and memory CD8+ T cell expansion and maintenance using various common γc cytokine treatments; to investigate the application of IL-7 and IL-21 to antigen-specific CD8+ T cell cloning via pentamer- and activation-marker-guided FACS, and to develop a protocol for the activation-marker-guided isolation and expansion of protein- or synthetic long peptide-specific CD4+ and CD8+ T cell clones without prior knowledge of their minimal peptide epitope. This study demonstrated that both naive and memory T cells could be isolated and cloned with high efficiency under the influence of IL-7 and IL-21, and retained proliferative potential and memory phenotype associated with anti-tumour efficacy in clinical trials. This study further demonstrated that temporal detection of cell surface activation markers allowed isolation of T cells of unknown epitope specificity, and facilitated the definition of novel peptide:MHC restrictions in T cells specific for the therapeutically relevant cancer-testis antigen NY-ESO-1.Items in ResearchSpace are protected by copyright, with all rights reserved, unless otherwise indicated. Previously published items are made available in accordance with the copyright policy of the publisher.https://researchspace.auckland.ac.nz/docs/uoa-docs/rights.htmhttp://creativecommons.org/licenses/by-nc-nd/3.0/nz/ThesisCopyright: The authorQ112931773