The Live Cell Imaging Shared Resource (LCISR) of Simmons Comprehensive Cancer Center (SCCC) enables SCCC members to leverage advanced light microscopy techniques in support of their cancer-focused research. The LCISR’s capabilities include digital imaging of histologically stained samples, workhorse confocal microscopes, sophisticated two-photon microscopes for imaging in live animals and thick tissues, total internal reflection fluorescence (TIRF) microscopes, and multiple modes of super-resolution imaging, as well as two image processing workstations with comprehensive image analysis software. Under the leadership of Kate Luby-Phelps, PhD, LCISR staff members provide individualized training of users and consult on experimental design and image analysis. The LCISR has been proactive in evaluating new instrumentation and in obtaining funding for instrumentation purchase. Since 2015, the LCISR has replaced two aging laser scanning confocal microscopes and has expanded its capacity by adding a third confocal with funding from the NIH S10 program. This acquisition included a Zeiss Airyscan FAST attachment for “super resolution” imaging and a fluorescence correlation spectroscopy module for examining molecular interactions in solution and in living cells that is unique on the UT Southwestern Medical Center (UTSW) campus. The LCISR recently acquired an OMX SR multimode super-resolution microscope with 3D structured illumination microscopy, localization microscopy (PALM/STORM), and Ring TIRF capabilities. To keep up with ongoing rapid developments in light microscope technology, the LCISR recently initiated a partnership with the UTSW Microscope Innovation Laboratory (MIL), led by Kevin Dean, PhD. The MIL is a newly established, institutionally supported initiative intended to fast-track the translation of cutting-edge light microscope imaging instrumentation to researchers at UTSW. Planned instrumentation includes custom-built lightsheet microscopes for cleared tissue imaging and fast volumetric imaging, and a three-photon microscope for deep tissue imaging. Instruments developed by the MIL will be made available to SCCC members through the LCISR. Since 2014, the LCISR has been used by more than 140 principal investigators across all five SCCC research programs, resulting in more than 140 cancer-focused publications, including publications in high-impact journals such as Cell, Nature, Science, Cancer Cell, Nature Communications, Dev Cell, and Nature Microbiology. The annual number of SCCC member laboratories using the LCISR has expanded from 54 in 2015 to 80 in 2019.