SUMMARY Membrane proteins play key roles in many cellular functions and are the largest class of drug targets. Measuring the molecular interactions of membrane proteins are critical for understanding protein functions, discovering biomarkers, and developing drugs. Most popular methods for measuring membrane protein interaction kinetics involves extraction, purification, stabilization of the membrane protein in an artificial lipid environment, which is not only labor intensive but may also introduce bias and costly missteps due to the loss of the heterogeneous native cellular microenvironment. Therefore, an in situ detection technique that enables sensitive measurement of native receptor behavior at single cell and population level is critical to expediting drug development. We propose to develop Plasmonic Scattering Microscopy (PSM) as a breakthrough technology for high throughput label-free quantification of membrane protein binding interaction kinetics on single cells. PSM advances the field of biomarker discovery and drug development by enabling high throughput real-time functional study of drug candidate interactions with cell membrane receptors in their native microenvironments. Surface plasmon resonance microscopy (SPRM) is the current state-of-the-are for studying label-free binding kinetics. PSM is a breakthrough advancement of SPR-based sensing. Rather than measuring changes in sensor reflectivity, PSM innovatively measures changes in plasmonic scattering. This novel approach enables quantitative real-time measurement of label-free molecular binding kinetics on single cells in high throughput but avoids the major drawbacks of reflection-based imaging known to plague traditional SPR-based approaches. Unlike SPRM, PSM exhibits several distinct technological advances, including a ~5 times greater signal to noise ratio, ~10 times greater field of view, ~5 times greater resolution, ~50 times higher throughput, enhanced sensitivity and data quality that avoids interferences from secondary reflection and near-field diffraction events, and the capability of simultaneous fluorescence imaging for orthogonal validation. In this fast-track STTR project, Biosensing Instrument Inc. (BI) will work with the inventor of PSM technology at Arizona State University to develop a commercial prototype multi-functional PSM instrument that can perform PSM and fluorescence imaging. We will also collaborate with potential customers in biomedical research and pharmaceutical industries to validate PSM performance and develop key applications. This project addresses the significant unmet need for acquiring more native biorelevant data sooner in the early-stage drug development process, thereby mitigating costly missteps and false leads. PSM technology enables sensitive label-free kinetic quantification of membrane protein behavior in their native cellular microenvironment with high throughput while also permitting simultaneous fluorescence validation. PSM addre...