NanoLuc® Luciferase Technology

NanoLuc® Luciferase (Nluc) is a 19.1 kDa, monomeric, ATP independent enzyme that utilizes a novel substrate to produce high intensity, glow-type luminescence. The enzyme was improved using directed evolution from a deep-sea shrimp luciferase, creating a luciferase that is much brighter than other forms of luciferase, including both firefly (Photinus pyralis) and Renilla reniformis. The high intensity luminescence of the NanoLuc enzyme combined with low autoluminescence of the furimazine substrate allows the sensitive detection of low levels of luciferase expression in experimental systems where other forms of luciferase can fail. Use NanoLuc Luciferase with the Nano-Glo™ family of assay reagents.

• Extremely Bright Output:

• Enhanced sensitivity allows detection of events at more physiologically relevant levels.
• In cells, signals average about 80x and can be up to 240x more than firefly luciferase (luc2)

• Incredibly Small Reporter (19kD)

• Much smaller than firefly or Renilla luciferases or GFPs
• Enables applications where gene or protein sizes are limiting, such as viral packaging or protein fusions.

• Glow Signal Kinetics:

• Generally suitable for all scales of laboratory throughput
• Readily adaptable to laboratory automation (e.g., HTS)

• High Physical Stability:

• Robust to experimental conditions (temperature, pH, etc.)
• Reduced false hits in compound library screening

• Highly Versatile:

• Standard assay format with cell lysis
• Secretion-based assays without lysis
• Intracellular assays with tunable protein stability

Relative sizes for different available luciferases.

Purified enzyme

Three different luciferases were measured for light output with equal molar amounts of enzyme (50 attomoles). Firefly luciferase was detected the ONE-Glo™ Luciferase Assay Reagent, Renilla luciferase with Renilla-Glo™ Luciferase Assay Reagent, and NanoLuc® luciferase with Nano-Glo™ Luciferase Assay Reagent on a GloMax® Multi Microplate Luminometer. It was found that NanoLuc® luciferase has an approximately 150-fold higher specific activity than the other luciferases.

Living HEK293 cells in 96-well plate  (50,000 cells per well). Imaged by a hand-held iPhone

NanoLuc® Luciferase is offered in a collection of 12 different vector configurations to best suit the intended use. These products offer: See All »

• Nluc gene sequence codon optimized and cleansed of most transcription factor binding sites, promoter modules, splice sites, and other regulatory sequences.
• Standard, destabilized, or secreted forms of luciferase available to match the most appropriate assay format.
• pGL4-based backbone for easy sequence transfer from existing plasmids
• Use a standard plasmid with Multiple Cloning Site (MCS) to clone promoters or Minimal Promoter-containing plasmid to clone response elements
• Hygromycin marker available for cell line creation
• CMV controls available in standard (intracellular) and secreted format
• NF-kB response element plasmid with destabilized NlucP for study of the NF-kB pathway

CRE response in HEK293 cells

Genetic reporter constructs where made using the indicated luciferase under control of the cAMP response element (CRE) and transfected into HEK293 cells. Following induction with forskolin, the luciferase expression was detected with the Nano-Glo™ or ONE-Glo® Luciferase Assay Systems on a GloMax® 96 Microplate Luminometer. NanoLuc® Luciferase demonstrates brighter signals than either standard or destabilized firefly luciferase. The PEST destabilization domain increases the induction ratio for both luciferases and provides maximal response when used with NanoLuc. In all cases the EC50 for the induction remains relatively unchanged.

HEK293 cells / CRE response element

The CRE-responsive expression of secreted NanoLuc® Luciferase (secNluc) was measured either in cells induced with forskolin or in DMSO-control over a period of 6 hours. A media change was performed at t=0. The secreted format allows the measurement of reporter activation kinetics without destroying the sample. Signals were measured with a GloMax® 96 Microplate Luminometer.

Nano-Glo™ Luciferase Assay System has been developed to provide optimal activity with the NanoLuc® Luciferase. This assay reagent offers:

• Add-Mix-read simplicity with integral lysis components
• Glow signal kinetics with an approximately 2 hour signal half-life
• Liquid substrate allows easy reagent preparation
• Minimal autoluminescense
• Room temperature stability through one work day
• Four sizes availabe

• 10ml
• 10x10ml
• 100ml
• 10x100ml