High Parameter Fluorescence Imaging for Analysis of Tissue Microenvironments - PROJECT SUMMARY AND ABSTRACT The Human Immune Monitoring Shared Resource (HIMSR) at the University of Colorado Anschutz Medical Campus (CUAMC) is applying for funds to add a state-of-the-art high-parameter fluorescence imaging platform, the Lunaphore COMET, to this well-established core facility. HIMSR is substantially supported by the University of Colorado Cancer Center (UCCC, P30 CA046934) and the Colorado Head and Neck Cancer SPORE (P50CA261605). Current multiplex immunohistochemistry (mIHC) tissue imaging instruments include the Akoya Vectra 3.0, the Akoya Vectra Polaris, and the Ionpath Multiplex Ion Beam Imaging (MIBI) system (low- throughput, 40-marker mass spectrometry imaging in regions of interest). HIMSR is the only core lab in the state of Colorado that offers multiplex tissue imaging and specializes in the development of customized high- parameter panels. HIMSR supports a consistently growing number of investigators, approximately 100 per year, 65% of which utilize mIHC instruments. The rationale for adding additional high-parameter imaging capability is that many investigators need affordable hyperplexing options, however the capacity of HIMSR's existing high- parameter MIBI platform is maximized with no available AUT for innovation or new assay development. The Lunaphore COMET instrument utilizes commercially available label-free primary antibodies in a serial staining, imaging, and elution approach that is flexible to each investigator's needs. It maximizes throughput with walk- away automation and preserves tissues and epitopes with unparalleled integrity and stability using innovative microfluidics, making it possible for investigators to perform whole genome RNA-based analyses of the same tissues in downstream transcriptomics assays. The Lunaphore COMET offers whole-tissue imaging with a virtually unlimited hyperplex capability and generates OME-TIFF image files that can be analyzed with either open-source or commercially available image analysis packages. Interest in high parameter tissue imaging and the need for additional instrumentation is driven by a strong group of NIH-funded principal investigators with 23 active NIH-funded grants in the areas of oncology, dermatology, hepatology, nephrology, and hematology. The estimated need for the 12 Major Users is approximately 81% accessible user time. The HIMSR team is highly qualified to develop new assays and operate the Lunaphore COMET instrument, composed of a master's level histologist, a Ph.D. level image analyst, a certified histotech, and three research assistants. The PI (Dr. Kimberly Jordan) has over 7 years of experience in developing, optimizing, and analyzing multiplex IHC tissue imaging data, has collaborated on many NIH-funded R01 projects utilizing these technologies and expertise, and authored 50 peer-reviewed publications in immunology. HIMSR's expertise in developing custom mIHC assays is nationally recognized with a strong track record of providing robust and quantifiable images of tissue microenvironments. There is a well-established financial and administrative infrastructure in place to support the Lunaphore COMET in HIMSR with strong leadership and institutional support.
