Red-Shifted Coumarin Luciferins for Improved Bioluminescence Imaging.

Multicomponent bioluminescence imaging requires an expanded collection of tissue-penetrant probes. Toward this end, we generated a new class of near-infrared (NIR) emitting coumarin luciferin analogues (CouLuc-3s). The scaffolds were easily accessed from commercially available dyes. Complementary mutant luciferases for the CouLuc-3 analogues were also identified. The brightest probes enabled sensitive imaging . The CouLuc-3 scaffolds are also orthogonal to popular bioluminescent reporters and can be used for multicomponent imaging applications. Collectively, this work showcases a new set of bioluminescent tools that can be readily implemented for multiplexed imaging in a variety of biological settings.