Functional Imaging Through Dark State Dynamics

This thesis harnesses the environmental sensitivity of the dark states of molecular fluorophores, both endogeneous in cells/tissue and externally introduced for mapping of chemical micro-environments including factors such as ion concentration and microviscosity. A novel technique for directly detecting the dynamics and population of dark states, such as the lowest triplet state, was developed and called FAPA, Fluorescence Anomalous Phase Advance. This technique is a sensitive and fast reporter of dark state dynamics and can be used for imaging. In addition, a genetically-encoded microviscosity and micro-environmental mapping ability with high protein-specific specificity was developed by using a TMP-tag and Fluorescence Lifetime Imaging.