Expanded palette of Nano-lanterns for real-time multicolor luminescence imaging

摘要: Fluorescence live imaging has become an essential methodology in modern cell biology. However, fluorescence requires excitation light, which can sometimes cause potential problems, such as autofluorescence, phototoxicity, and photobleaching. Furthermore, combined with recent optogenetic tools, the light illumination trigger their unintended activation. Because luminescence does not require it is a good candidate alternative modality to circumvent these problems. The application of imaging, however, been limited by two drawbacks existing luminescent protein probes, luciferases: namely, low brightness poor color variants. Here, we report development bright cyan orange proteins extending our previous yellowish-green Nano-lantern. change enhancement were both achieved bioluminescence resonance energy transfer (BRET) from enhanced Renilla luciferase fluorescent protein. Nano-lanterns was ∼20 times brighter than wild-type luciferase, allowed us perform multicolor intracellular submicron structures. rapid dynamics endosomes peroxisomes visualized at around 1-s temporal resolution, slow focal adhesions continuously imaged for longer few hours without photobleaching or photodamage. In addition, extended simultaneous monitoring multiple gene expression Ca2+ different cellular compartments single cell.