Our website does not fully support your browser.

We've detected that you are using an older version of Internet Explorer. Your commerce experience may be limited. Please update your browser to Internet Explorer 11 or above.

We believe this site might serve you best:

United States

United States

Language: English

Promega's Cookie Policy

Our website uses functional cookies that do not collect any personal information or track your browsing activity. When you select your country, you agree that we can place these functional cookies on your device.

Nominations are open for the Diversification Of Our Research Scientists Program! Nominate Now ›

Choosing a Luciferase Reporter Assay

A reporter assay comparison guide.

This guide will help you learn about reporter assays and find the one best suited for your needs. Visit the Promega catalog listing to view ordering information for the Reporter Assays featured here.

Reporter Genes and Assay Chemistries

When creating a luciferase reporter assay, there are two important elements to consider: the luciferase reporter protein itself and the assay chemistry used to detect reporter activity. The characteristics of these two components together contribute to the overall performance of the assay. By selecting the reporter/detection solution that is optimal for your experimental goals, you can customize your luciferase reporter assay to create the best solution for your research.

Promega offers a choice of 3 different luciferase reporters: NanoLuc® Luciferase(Nluc,19kDa), Renilla Luciferase (Rluc, 36kDa) and Firefly Luciferase (Fluc, 61kDa) which vary in size, brightness, and protein half-life.

Luciferase Reporters Reporter gene names
Firefly Luciferase Firefly Luciferase descriptive text
luc, luc+, luc2, luc2P, luc2CP

Luc/Luc+: Original luciferase gene used in older luciferase reporter systems.

Luc2: Codon-optimized version of luciferase, engineered to remove most cryptic transcription factor binding sites and improve mammalian expression.

Luc2P and Luc2CP: Destabilized versions of Luc2 containing the PEST protein degradation sequence for rapid response.

Renilla Luciferase
Rluc, hRluc, hRlucP, hRlucCP

Rluc: Renilla luciferase.

hRluc: Synthetic codon-optimizied Renilla luciferase, engineered to remove cryptic transcription factor binding sites for improved mammalian expression.

hRlucP and hRlucCP: Destabilized versión of hRluc containing the PEST protein degradation sequence for rapid response.

NanoLuc Luciferase
Nluc, NlucP, secNluc

Nluc: NanoLuc luciferase.

NlucP: Nanoluc luciferase with PEST protein degradation sequence for rapid response.

secNluc: Secreted NanoLuc luciferase.

Detection Reagent Considerations

Multiple assay detection reagents are also available for each reporter. Key considerations for selecting the optimal assay reagent include:

  • Signal intensity and overall dynamic range needed for the assay.
  • Signal stability, or half-life, which will impact your processing workflow.
  • Processing steps required. Non-homogenous assays require a separate lysate creation step prior to reagent addition. Homogenous assay reagents are added directly to the cells in culture eliminating sample pre-processing.
  • Lytic or live-cell reporter detection
  • Single or dual-reporter detection

Luciferase Signal Strength and Stability

Dual-Luciferase® Assay Options

Firefly Luciferase Assay Options

RECOVER_RECOVER_12881mb-animation_HTML5 Canvas

Here, we compared luminescence signals from HEK293 cells transfected with a 1:1:8 ratio of either TK-Rluc (Renilla):TK-Fluc (firefly):carrier DNA or TK-Nluc (NanoLuc):TK-Fluc:carrier DNA and assayed with NanoDLR™, DLR™ or Dual-Glo® Luciferase Assay Systems as indicated. The NanoDLR(TM) and Dual-Glo(R) reagents are homogenous assay systems with increased signal stability. The DLR(TM) reagent is a non-homogenous assay with a flash signal that decays rapidly.

Here we demonstrate luminescence signal over time from a dilution of QuantiLum® Recombinant Luciferase assayed with various firefly luciferase detection reagents as indicated. The Luciferase Assay System is a non-homogenous reagent that provides the brightest initial luminescence with flash kinetics that has rapid signal decay. The Bright-Glo™, ONE-Glo™, ONE-Glo™ EX, and Steady-Glo® systems are homogenous reagents that show progressively decreasing levels of initial brightness with respective increases in signal half-life.

Luciferase Signal Strength and Stability

Dual-Luciferase® Assay Options

12881MB-W

Here, we compared luminescence signals from HEK293 cells transfected with a 1:1:8 ratio of either TK-Rluc (Renilla):TK-Fluc (firefly):carrier DNA or TK-Nluc (NanoLuc):TK-Fluc:carrier DNA and assayed with NanoDLR™, DLR™ or Dual-Glo® Luciferase Assay Systems as indicated. The NanoDLR(TM) and Dual-Glo(R) reagents are homogenous assay systems with increased signal stability. The DLR(TM) reagent is a non-homogenous assay with a flash signal that decays rapidly.

Firefly Luciferase Assay Options

12881mb-w3

Here we demonstrate luminescence signal over time from a dilution of QuantiLum® Recombinant Luciferase assayed with various firefly luciferase detection reagents as indicated. The Luciferase Assay System is a non-homogenous reagent that provides the brightest initial luminescence with flash kinetics that has rapid signal decay. The Bright-Glo™, ONE-Glo™, ONE-Glo™ EX, and Steady-Glo® systems are homogenous reagents that show progressively decreasing levels of initial brightness with respective increases in signal half-life.


Compare Luciferase Assay Characteristics

Search by luciferase or assay type to compare options

Luciferase Detected Assay Reagent Ideal For Signal Half-Life Sensitivity Number of Steps Live Cell Assay? Injectors Needed?
NanoLuc, Firefly Nano-Glo® Dual-Luciferase® Reporter Assay System (NanoDLR) Homogenous Fluc/Nluc dual-reporter detection with flexibility in choice of primary reporter. Provides highest sensitivity for both reporters when a stable signal is required and highest sensitivity option when Nluc is used as the primary reporter. Ideal for low to high-throughput processing when 2 primary reporters or internal control is needed. 2 hours each +++(Fluc)
+++++(Nluc)
2
(lysate optional)
No Optional
Firefly,
Renilla
Dual-Luciferase® Reporter Assay System (DLR) Non-homogenous Fluc/Rluc dual-reporter detection. Requires lysate creation and 2 injectors for delivery. Ideal for small sample numbers when maximum Fluc sensitivity and internal control are needed. 10 minutes, 2 minutes ++++(Fluc)
++++(Rluc)
3 No Yes
2 injectors
Firefly,
Renilla
Dual-Glo® Luciferase Assay System Homogenous Fluc/Rluc dual-reporter detection with reduced sensitivity and stable signal. Ideal for high-throughput processing of Fluc reporter when Rluc is used as the internal control. 2 hours each ++
(Fluc/Rluc)
2 No No
NanoLuc Nano-Glo® Luciferase Assay System Homogenous Nluc detection with bright, stable signal. Ideal for low to high-throughput processing when maximum sensitivity is required. 2 hours +++++ 1 No, unless used with secNluc No
NanoLuc Nano-Glo® Live Cell Assay System Live cell Nluc detection with brightest signal. Ideal for single timepoint analysis when high sensitivity is needed in a non-lytic assay. 30 minutes ++++ 1 Yes No
NanoLuc Nano-Glo® Vivizine™ Live Cell Substrate Live cell Nluc detection with intermediate signal and stability. Ideal for kinetic analysis lasting multiple hours. 12 hours +++ 1 Yes No
NanoLuc Nano-Glo® Endurazine™ Live Cell Substrate Live cell Nluc detection with most stable signal. Ideal for multi-day kinetic analysis. Up to 72 hours ++ 1 Yes No
Firefly Luciferase Assay System Non-homogenous Fluc detection requiring lysate creation and injector delivery. Ideal for small sample numbers when maximum Fluc sensitivity is needed. 10 minutes ++++ 2 No Yes
Firefly Bright-Glo® Luciferase Assay System Homogenous Fluc detection offering the brightest signal and shortest half-life. Ideal for high-throughput processing when high sensitivity is required. 30 minutes +++ 1 No No
Firefly ONE-Glo™ Luciferase Assay System Homogenous Fluc detection with moderate signal and half-life. Ideal for high- or ultrahigh-throughput processing. 45 minutes ++ 1 No No
Firefly ONE-Glo™ Ex Luciferase Assay System Homogenous Fluc detection with moderate signal and half-life and improved storage stability. Ideal for high- or ultrahigh-throughput processing and repeat use. Also used to detect Fluc in NanoDLR allowing consistent use between single and dual assays. 2 hours ++ 1 No No
Firefly Steady-Glo® Luciferase Assay System Homogenous Fluc detection with maximum signal stability and reduced signal. Ideal for high-throughput applications when extended luminescence is required. 5 hours + 1 No No
Renilla Renilla Luciferase Assay System Non-homogenous Rluc detection requiring lysate creation and injector delivery. Ideal for small sample numbers when maximum Rluc sensitivity is needed. 2 minutes ++++ 2 No Yes
Renilla Renilla-Glo Luciferase Assay System Homogenous Rluc detection with increased signal stability. Ideal for high-throughput processing. 40+ minutes ++ 1 No No
Renilla ViviRen™ Live Cell Substrate Live cell Rluc detection with highest signal. Ideal for single time point or short non-lytic analysis when highest sensitivity is required. 10 minutes +++ 1 Yes No
Renilla EnduRen™ Live Cell Substrate Live cell Rluc detection with greatest stability. Ideal for extended kinetic analysis. Up to 24 hours + 1 Yes No

Learn More About Bioluminescent Reporter Assay Design

This two-part series begins by walking through the basic considerations for choosing the optimal experimental reporter. In part two, Assay Normalization Options", experimental design and data analysis methods are discussed.