@article{eace26bd16a74bbe9e7e93762a13a662,
title = "A bright cyan-excitable orange fluorescent protein facilitates dual-emission microscopy and enhances bioluminescence imaging in vivo",
abstract = "Orange-red fluorescent proteins (FPs) are widely used in biomedical research for multiplexed epifluorescence microscopy with GFP-based probes, but their different excitation requirements make multiplexing with new advanced microscopy methods difficult. Separately, orange-red FPs are useful for deep-tissue imaging in mammals owing to the relative tissue transmissibility of orange-red light, but their dependence on illumination limits their sensitivity as reporters in deep tissues. Here we describe CyOFP1, a bright, engineered, orange-red FP that is excitable by cyan light. We show that CyOFP1 enables single-excitation multiplexed imaging with GFP-based probes in single-photon and two-photon microscopy, including time-lapse imaging in light-sheet systems. CyOFP1 also serves as an efficient acceptor for resonance energy transfer from the highly catalytic blue-emitting luciferase NanoLuc. An optimized fusion of CyOFP1 and NanoLuc, called Antares, functions as a highly sensitive bioluminescent reporter in vivo, producing substantially brighter signals from deep tissues than firefly luciferase and other bioluminescent proteins.",
author = "Jun Chu and Younghee Oh and Alex Sens and Niloufar Ataie and Hod Dana and Macklin, {John J.} and Tal Laviv and Welf, {Erik S.} and Dean, {Kevin M.} and Feijie Zhang and Kim, {Benjamin B.} and Tang, {Clement Tran} and Michelle Hu and Baird, {Michelle A.} and Davidson, {Michael W.} and Kay, {Mark A.} and Reto Fiolka and Ryohei Yasuda and Kim, {Douglas S.} and Ng, {Ho Leung} and Lin, {Michael Z.}",
note = "Funding Information: We thank T. Doyle (Stanford University) for assistance with bioluminescence imaging, J. Chen (Shenzhen Institutes of Advanced Technology) for analysis of light-sheet microscopy data, S. Classen (ALS SIBYLS) for help with synchrotron X-ray data collection, P. Meisenheimer and T. Kirkland (Promega) for furimazine, and members of the Lin laboratory for general assistance and advice. The MV3 cells were a gift of P. Friedl, University of Texas MD Anderson Cancer Center. The Advanced Light Source (ALS), a national user facility operated by Lawrence Berkeley National Laboratory on behalf of the Department of Energy Office of Basic Energy Sciences, through the Integrated Diffraction Analysis Technologies (IDAT) program, is supported by the DOE Office of Biological and Environmental Research and National Institute of Health project MINOS (R01GM105404). This work was supported by the University of Hawaii at Manoa Undergraduate Research Opportunities Program (M.H. and C.T.T.), a long-term fellowship from the Human Frontier Science Program (T.L.), NIH grants R01HL064274 (M.K.), R01MH080047 (R.Y.), 1U01NS090600 (M.Z.L.), and P50GM107615 (M.Z.L.), Shenzhen Basic Research Foundation grant JCYJ20150521144320987 (J.C.), the Hundred Talents Program award (Y64401) from the Chinese Academy of Sciences (J.C.), a Burroughs Wellcome Foundation Career Award for Medical Scientists (M.Z.L.), and a Rita Allen Foundation Scholar Award (M.Z.L.).",
year = "2016",
month = jul,
day = "1",
doi = "10.1038/nbt.3550",
language = "אנגלית",
volume = "34",
pages = "760--767",
journal = "Nature Biotechnology",
issn = "1087-0156",
publisher = "Nature Research",
number = "7",
}