Summary (1R35GM134963). Understanding the mechanisms of protein and organelle function and dysfunction requires dynamic analysis of molecular structures and interactions in live cells. A long-term goal of my laboratory is to develop tools that reveal these functional elements and apply them to discover novel a significant biology and molecules with useful or influential properties. I am drawn to complex, multi-disciplinary problems where chemistry catalyzes exciting and non-incremental progress. We strive not simply to develop tools but also apply them in innovative ways. Fluorescence spectroscopy features prominently in the two research programs described herein. In both, fluorescent tools that we developed using prior NIGMS support have uncovered influential new biology; this application continues this development and expands the new biology to benefit human health.