FLOW CYTOMETRY CORE

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Core A contains a Becton-Dickinson DiVa cell sorter and LSR analyzer. The DiVa is a 13-parameter 4-way 5-laser cell sorter. The LSR has excitation lines in the ultraviolet, visible and red spectra. The Core Leader was recently awarded (1/1/05) a SIG to build a 6-laser, 20-detector LSR-2 which will be available for this program. For Project 1 (Sykes), Core A will be used for 9-color flow cytometry studies required to assess activation and memory markers on GVH-reactive and non-reactive T cell subsets identified with specific markers, for CFSE studies, and for sorting of T cell subsets to be adoptively transferred from lymphoid and non-lymphoid tissues of GVHD mice and delayed DLI recipients. Project 2 (Yang) will investigate the mechanisms by which IFN-gamma separates GVL effects from GVHD-inducing activity of donor CDS T cells in allogeneic HCT recipients. The program will need a cell sorter about 2 days/month to purify donor from recipient mouse cells, obtained from approximately 30 mice/yr, for transplantation. For Project 3 (Sachs), Core A will be used for the specific T-cell subset depletion of the donor DLI inoculum and the separation of donor-type and host-type T-cells from PBMC of chimeric recipients at monthly intervals, following HCT for the purpose of in vitro and in vivo mechanistic studies of regulatory cells. For Project 4 (Scadden), the result of PTH administration on CD34+ cell yields in patients treated with G-CSF mobilization will be monitored. The immune reconstitution of post-transplant patients treated with PTH will also be monitored. For the murine studies related to Project 4, Core A will assess and separate lymphocytes based upon T-cell differentiation antigens and MHC specificity. Approximately 20 sorts and 45 analyses will be needed for this project.