FLOW CYTOMETRY & CELL SORTING SHARED RESOURCE: PROJECT SUMMARY Flow cytometry is a mainstay technology required to support a dynamic range of basic science and clinical biomedical research programs across scientific disciplines. The Flow Cytometry & Cell Sorting Shared Resource (FCSR) provides an array of options for both (1) cell analysis and (2) cell sorting required for functional cellular analysis or downstream molecular inquiry. Due to the nature of FCSR services, flexible on-site flow cytometry facilities are vital to maintain the viability of sorted cells and to perform time-sensitive analysis that supported 30 Georgetown Lombardi Comprehensive Cancer Center (LCCC) Members in FY2022. Over the current project period the FCSR has been transformed into a multi-site Shared Resource (SR), which now provides NJ-based LCCC Members access to on-site FCSR services. Led by Director Karen Creswell, PhD and Manager Yuanyuan Tian, PhD, who oversees FCSR operations on the NJ campus, the FCSR support staff (three FTEs) provide flow cytometry services, experimental and panel design, and post-acquisition analysis. The development of new methodologies is also prioritized to ensure researchers have access to the most current flow cytometry services. Creswell and Tian have integrated and standardized FCSR services across the Consortium. The FCSR provides value to research projects by separating cells based on transfection-based markers, stem cell panels, and additional cell surface and intracellular markers. Functional assays for cell cycle, apoptotic status, and oxidative stress are performed in addition to immunophenotyping. Immunophenotyping of mouse and human tissues and tumor infiltrating lymphocytes (TILs) has improved due to the 16-color capabilities of the BD LSRFortessa and the 28-color capabilities of the BD FACSymphony A3 cell analyzer. The FCSR receives input from the FCSR Advisory Committee (AC) and annual user surveys to prioritize development and assess programmatic needs across the consortium. To this end, new FCSR initiatives aim to provide cutting-edge mass cytometry access to LCCC Members through the addition of the Standard BioTools CyTOF XT system, greatly expanding user analysis to more than 40 simultaneous markers, providing an invaluable tool for cancer and immune studies. In FY2022, the FCSR provided services to 30 LCCC Members across two Research Programs (12 Cancer Cell Biology [CCB] members, and 18 Cancer Host Interactions [CHI] members). During the current project period, the FCSR contributed to 52 publications, of these 28 in high impact (IF>10) journals including Nature, Nat Immunol., and Cell Mol Immunol. The FCSR also provided support that enabled the success of three clinical trials and 36 extramurally-funded research project grants.