Abstract The goal of this proposal is to secure funds to acquire a Zeiss Crossbeam 550 Field Emission Scanning Electron Microscope (SEM) with an Ion-sculptor Focused Ion Beam (FIB) column and a Quorum PP3010Z cryo-stage to support NIH-funded research efforts at Emory University and member institutions of the Georgia Core Facilities Partnership (GCFP). The cryo-FIB-SEM instrumentation will allow researchers to carry out three-dimensional (3D) analysis of biological samples under room temperature and cryogenic conditions. This equipment will be used to investigate cellular ultrastructure in 3D with enough resolution to observe details of cellular organelles and generate quantitative data. The dual beam system allows for multiple sample types to be processed and for electron tomography (ET) data to be obtained in situ, or through downstream cryo-ET of FIB-milled lamella. This equipment will have direct applications in disease biology research, benefiting projects investigating infectious diseases (Melikian, Sarafianos, Liang, Neufeldt, and D’Souza), neurodegenerative disorders (Smith, Kukar, and D’Souza), heart conditions (Kwong), liver diseases (Yang), cancer (Yang, Liu, and Moniri), and the development of therapeutics and diagnosis systems (Sarafianos, Yang, Banga, and Chougule). The instrument will be housed and administered by the Robert P. Apkarian Integrated Electron Microscopy Core (IEMC), with an established infrastructure for equipment administration, maintenance, and management, and an exceptional staff to support research and train investigators in sample preparation, data acquisition, and processing. In summary, acquisition of a cryo-FIB-SEM will offer imaging capabilities currently unavailable for biomedical research at Emory and other regional institutions. It will have an immediate and profound impact on their projects by allowing image acquisition on site, direct feedback from researchers, training of junior and senior investigators in FIB-SEM applications, rapid turnaround to produce results for publications and grants, and a reduction in research costs by removing the need for outsourcing.