Pierce™ Firefly Luciferase Glow Assay Kit
Pierce™ Firefly Luciferase Glow Assay Kit
Thermo Scientific™

Pierce™ Firefly Luciferase Glow Assay Kit

The Thermo Scientific Pierce Firefly Luciferase Glow Assay Kit provides a bright and stable bioluminescence signal (half-life greater than 2Read more
Have Questions?
Change viewbuttonViewtableView
Catalog NumberQuantity
16176100 Reaction Kit
161771000 Reaction Kit
Catalog number 16176
Price (USD)
134.00
Each
Add to cart
Quantity:
100 Reaction Kit
Price (USD)
134.00
Each
Add to cart
The Thermo Scientific Pierce Firefly Luciferase Glow Assay Kit provides a bright and stable bioluminescence signal (half-life greater than 2 hours) in the presence of firefly luciferase reporters for protein expression assays.

Features of the Firefly Luciferase Glow Assay Kit:

Sensitive—highly sensitive detection of firefly luciferase activity; for best results (maximum signal) use with Pierce Firefly Luciferase Signal Enhancer
Stable—increased signal stability compared to flash assays
Simple—does not require a luminometer with injectors
Compatible—assay reagents compatible with other firefly beetle luciferases
Automation-friendly—amenable to high throughput assays
Convenient—contains a universal cell lysis buffer and optimized glow assay reagent
Safe—allows one to perform non-radioactive assays

This Pierce Luciferase Glow Assay Kit contains reagents for measuring the activity of firefly luciferase in cell lysates. In the presence of firefly luciferase, the glow reagent provides stable bioluminescent signal and is highly suited for luminometers without injectors or for batch processing of samples. The light output generated by the luciferase reaction can be correlated with the amount of luciferase protein produced which in turn is proportional to the promoter activity driving the luciferase expression. The glow assay reagents were optimized for best results with Thermo Scientific Firefly Luc Plasmids; however, this assay kit may be used to detect the activity of other firefly or ATP-dependent luciferases that use D-luciferin as a substrate.

Includes:
Cell lysis buffer, reaction buffer and substrate; purchase Luciferase Signal Enhancer separately.

Requires:
• Firefly luciferase and luminometer or other instrument capable of monitoring luminescence, such as Thermo Scientific Luminoskan Ascent and Varioskan Flash Microplate Readers.
• For best results (maximum signal), use with Pierce Firefly Luciferase Signal Enhancer

Applications:
• Promoter studies for analyzing cis- regulatory elements and trans-acting factors
• Drug screening
• siRNA and miRNA screening
• Multiplexed assays to study off-target effects
• Protein localization reporter assays
• Signal transduction pathway analysis
• RNA splicing studies

Firefly luciferase is a 60kDa protein produced in nature by several species of the Lampyridae family of beetles which includes the genera Photinus and Luciola. Bioluminescent signal from firefly luciferase originates from the oxidation of D-luciferin. Light output generated by the luciferase reaction is captured using a luminometer and can be correlated with the amount of firefly luciferase protein produced and used to determine the activity of the promoter driving firefly expression. The native firefly luciferase produces a strong flash signal that decays rapidly. The signal produced using the Pierce Firefly Glow Assay Kit is more stable and decays very slowly, thereby eliminating the need for a luminometer with injectors.
For Research Use Only. Not for use in diagnostic procedures.
Specifications
TypeAssay Kit
TargetLuciferase, Firefly Luciferase
Quantity100 Reaction Kit
AssayReporter Enzyme, Luciferase Reporter Assay
Compatible CellsMammalian Cells
Detection MethodBioluminescence
For Use With (Equipment)Luminometer (Microplate)
Format384-well plate, 96-well plate
Label TypeEnzyme Labeled
Substrate PropertiesChemical Substrate
TechniqueEnhanced Chemiluminescence
Substrate TypeLuciferase Substrate
SubstrateD-Luciferin
Unit SizeEach
Contents & Storage
• Firefly Glow Assay Buffer, 5 mL, store at -20°C
• D-Luciferin, Lyophilized, 3 mg, store at 4°C
• 2X Cell Lysis Buffer, 6 mL, store at room temperature

Upon receipt store kit at -20°C or store individual components as indicated above.

Frequently asked questions (FAQs)

I got a high background signal with a Firefly Luciferase Glow/Flash assay. Why is this?

This could be due to contamination of the control sample. Make sure to use new sample and change pipette tips after each well.

Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.

I got a high signal with a Firefly Luciferase Glow/Flash assay. What could have happened?

This could be due to high luciferase expression. Here are some suggestions:

  • Reduce incubation time before collecting samples. 
  • Decrease the integration time on the instrument.
  • Dilute the sample 
    Note: A low sample volume can increase assay variability. Dilute the sample and use the recommended volume of 10-20 µL per assay



  • Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.

    What could be causing no signal or low signal with the Firefly Luciferase Glow/Luciferase Flash assay? Do you have any suggestions to get around this problem?

    Here are possible causes and solutions:

    - Low transfection efficiency: Optimize transfection conditions using a visual transfection control (e.g., a plasmid over-expressing a fluorescent protein); Verify plasmid DNA quality - use only transfection grade DNA; Use actively dividing, low passage cells; Use a different cell type.
    - No or low promoter activity: Use conditions known for promoter activation; Incubate cells for a longer time; Change growth conditions to improve expression; Use a different promoter.
    - D-luciferin auto-oxidized: Protect substrate from light and maintain 100X D-luciferin at -20 degrees C; Prepare new Working Solution if used longer than 4 hours.
    - Low luciferase expression: Use Pierce Firefly Signal Enhancer (100X) (Cat. No. 16180); Lyse cells in a smaller volume of 1X Cell Lysis Buffer; Use a different promoter or growth conditions to improve expression; Increase the integration time on the instrument; Scale-up the volume of sample and reagent per well.
    - Degraded luciferase protein: Store cell lysates on ice and perform assays immediately following cell lysis.

    Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.

    I got a high background signal with a Gaussia/Cypridina/Renilla Luciferase Glow/Luciferase Flash assay. What went wrong?

    Here are possible causes and solutions:

    - Non-specific oxidation of substrate: Use less serum in the cell culture media; Note: Albumin can increase the auto-oxidation of Vargulin; Avoid repeated freezing and thawing of the sample.
    - Control sample is contaminated: Use new sample; Change pipette tips after each well.

    Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.

    I am getting a high saturating signal with a Gaussia/Cypridina/Renilla Luciferase Glow/Luciferase Flash assay. What could have happened?

    This could be due to high luciferase expression. Here are some suggestions:

  • Reduce incubation time before collecting samples. 
  • Decrease the integration time on the instrument.
  • Dilute the sample: for secreted Gaussia/Cypridina Luciferase, dilute the sample using media from the cell culture; for cell lysate, dilute the sample using lysis buffer 


  • Note: A low sample volume can increase assay variability. Dilute the sample and use the recommended volume of 10-20 µL per assay.

    Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.