2. Academic Validation
  3. Novel NanoLuc substrates enable bright two-population bioluminescence imaging in animals

Novel NanoLuc substrates enable bright two-population bioluminescence imaging in animals

  • Nat Methods. 2020 Aug;17(8):852-860. doi: 10.1038/s41592-020-0889-6.
Yichi Su  # 1 2,  Joel R Walker  # 3,  Yunhee Park 1 2,  Thomas P Smith 3,  Lan Xiang Liu 1 2,  Mary P Hall 4,  Louai Labanieh 2,  Robin Hurst 4,  David C Wang 1 5,  Lance P Encell 4,  Namdoo Kim 1 2 6,  Feijie Zhang 7 8,  Mark A Kay 7 8,  Kerriann M Casey 9,  Robbie G Majzner 7 10,  Jennifer R Cochran 2,  Crystal L Mackall 7 11,  Thomas A Kirkland 12,  Michael Z Lin 13 14 15
Affiliations

Affiliations

  • 1 Department of Neurobiology, Stanford University, Stanford, CA, USA.
  • 2 Department of Bioengineering, Stanford University, Stanford, CA, USA.
  • 3 Promega Biosciences LLC, San Luis Obispo, CA, USA.
  • 4 Promega Corporation, Madison, WI, USA.
  • 5 Department of Biology, Stanford University, Stanford, CA, USA.
  • 6 Department of Chemistry, Kongju National University, Gongju, South Korea.
  • 7 Department of Pediatrics, Stanford University, Stanford, CA, USA.
  • 8 Department of Genetics, Stanford University, Stanford, CA, USA.
  • 9 Department of Comparative Medicine, Stanford University, Stanford, CA, USA.
  • 10 Stanford Cancer Institute, Stanford University, Stanford, CA, USA.
  • 11 Department of Medicine, Stanford University, Stanford, CA, USA.
  • 12 Promega Biosciences LLC, San Luis Obispo, CA, USA. thomas.kirkland@promega.com.
  • 13 Department of Neurobiology, Stanford University, Stanford, CA, USA. mzlin@stanford.edu.
  • 14 Department of Bioengineering, Stanford University, Stanford, CA, USA. mzlin@stanford.edu.
  • 15 Department of Pediatrics, Stanford University, Stanford, CA, USA. mzlin@stanford.edu.
  • # Contributed equally.
PMID: 32661427 DOI: 10.1038/s41592-020-0889-6
Abstract

Sensitive detection of two biological events in vivo has long been a goal in bioluminescence imaging. Antares, a fusion of the luciferase NanoLuc to the orange fluorescent protein CyOFP, has emerged as a bright bioluminescent reporter with orthogonal substrate specificity to firefly luciferase (FLuc) and its derivatives such as AkaLuc. However, the brightness of Antares in mice is limited by the poor solubility and bioavailability of the NanoLuc substrate furimazine. Here, we report a new substrate, hydrofurimazine, whose enhanced aqueous solubility allows delivery of higher doses to mice. In the liver, Antares with hydrofurimazine exhibited similar brightness to AkaLuc with its substrate AkaLumine. Further chemical exploration generated a second substrate, fluorofurimazine, with even higher brightness in vivo. We used Antares with fluorofurimazine to track tumor size and AkaLuc with AkaLumine to visualize CAR-T cells within the same mice, demonstrating the ability to perform two-population imaging with these two luciferase systems.

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