Development and Characterization of Novel Bioluminescent Reporters of Cellular Activity

Abstract

Over the years, periodic innovations in fluorescence microscopic technologies have afforded researchers a more sophisticated understanding of the inner workings of the cell. These methods are valuable but drawbacks include phototoxicity, photobleaching, low signal-to-noise ratio, and high background intensities distracting and sometimes disrupting the phenomena being studied. To circumvent these issues, we designed proof-of-principle bioluminescent sensors and applied them to common problems scientists face while using fluorescence imaging. Bright, bioluminescence-based, ratiometric Ca2+ sensors CalfluxVTN and CalfluxCTN can track intracellular Ca2+ concentration and can be used in conjunction with optogenetic stimulation in scenarios when fluorescence excitation illumination would be problematic. Additionally, Calflux sensors can be optimally applied to high throughput screening assays where fluorescent drug-like compounds are being screened all while taking advantage of already available microscopic technologies.