PROJECT SUMMARY/ABSTRACT The Sidney Kimmel Comprehensive Cancer Center (SKCCC) Flow/Mass Cytometry Shared Resource (SR) has continuously offered state-of-the-art cell sorting and flow analysis services to SKCCC Members from its inception in 1980. In 2013, the Flow Cytometry SR merged with the Human Immunology (HI) SR, but HI has since been more appropriately integrated with a newly established SR that handles clinical specimen collections and processing for all SKCCC studies. To expand the capabilities for tumor microenvironment and immune profiling, the SKCCC recently acquired mass cytometry technologies for suspension and imaging analyses. These resources were integrated into the collective Flow/Mass Cytometry and Technology Development (FCTD) Shared Resource (SR) in 2020. While many diverse therapies are being evaluated at the SKCCC, many studies overlap in their requirement for flow/mass cytometry analysis and cell sorting of immune cells. These services are provided in a timely, flexible and affordable manner to SKCCC Members from across the medical campus. Services provided by the FCTD SR do not duplicate services available elsewhere, and the SR currently operates the only mass cytometer at the institution. Cytometry services range from basic two- color flow analyses to complex cell-sorting experiments involving up to 16 colors, as well as 20-plex mass cytometry profiling based on 30-40+ parameters, plus training, data analysis and consultation services. Successful execution of these services has led to: 1) a novel analytical pipeline for determining T cell repertoire changes in response to immunotherapy regimens (i.e., the Mutation-Associated Neoantigen Functional Expansion of Specific T Cells Assay), 2) synergistic incorporation of multimer-based methodologies for investigating neoantigen-specific T cell subpopulations, and 3) a platform for innovative integration of scRNAseq datasets to study T cell responses in collaboration with the SKCCC Experimental and Computational Genomics SR. Moreover, the application of mass cytometry techniques, as currently offered by the FCTD SR, has already enabled discovery for preclinical and translational researchers at the SKCCC. Collectively, these resources significantly improve the ability to functionally characterize immune cells in cancer therapy, vaccine trials and preclinical studies. SKCCC Managed Shared Resource Reporting Period: January 1, 2020 to December 31, 2020