Project Summary Our goal is to develop a fluorescence guided imaging solution to lung cancer. Each year, lung cancer is the biggest cause of cancer related deaths in the US (>160,000 people) and the world (1.4 million fatalities). The primary strategy in this Project is to develop, characterize and test a combination of four targeted near-infrared contrast agents that will be used in humans to improve procedures for patients with lung cancer. It is clear that using fluorescent targeted agents during cancer surgery can highlight cancer cells for clinicians, thus several groups are testing molecular probes. Our research will specifically focus on lung cancer. Lung cancer is a very heterogeneous tumor due to the wide range of offending agents that can cause this disease. The proposed work will optimize four distinct types of contrast agents in a combination or cocktail for a translational intraoperative imaging clinical trials. The first cocktail agents are composed of folate and GCPII conjugated to a NIR dye. These two contrast agents are fully functional and are in human trial. However, together, they still miss a significant proportion of patients with lung cancer. Thus, we will be focusing our efforts on two more agents, CAIX and FAP, in this proposal. The goal of this Aim is to integrate the NIR tracers that are ready for clinical use. We will be utilizing both murine models and spontaneous lung cancer models in pet canines. Integration of these four agents will require studying the changes in sensitivity and specificity. It will also require examining the timing of these agents and the interactions of these tracers. If this cocktail is successful, it will provide a complete solution for the number one cancer killer in the United States.