In vivo animal imaging is an outstanding noninvasive tool to study the pathophysiology of disease or response to therapy; additionally, serial imaging reduces the required number of experimental animals. Because of the tremendous capital investment, we recommend the imaging center be a shared resource to facilitate innovative and productive cross-disciplinary scientific collaborations. A shared center also enables a broader range of imaging, as equipment is often cost prohibitive for smaller facilities. A multitude of factors will determine the architectural design, facility efficiency, and functionality. Important considerations to determine during the planning stages include the types of animals to be imaged, types of imaging studies to be performed, types of imaging equipment and related services to be offered, and the location of the imaging center. Architects must work closely with manufacturers to accommodate equipment-related building specifications; facility planners and veterinarians can provide a practical logistical design that will ensure efficient functionality. Miscellaneous considerations include biosecurity levels, use of radioisotopes, and personnel safety in the imaging environment. The ideal imaging center will include space to house animals and perform necessary preimaging procedures, state-of-the-art in vivo imaging devices and the most up-to-date anesthesia, physiological support, and monitoring equipment. The center staff should include imaging specialists for technical development and data analysis. As it is difficult to provide a comprehensive manual for setting up an in vivo animal imaging center, we offer advice based on our experiences with the National Institutes of Health Mouse Imaging Facility. Because magnetic resonance imaging (MRI) is the most expensive imaging tool, requires specific building design considerations, and poses unique occupational health and safety risks, we focus on MRI as the foundation for an imaging facility design.