Amplite™ Renilla Luciferase Reporter Gene Assay Kit *Bright Glow*

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1e+51e+410001001010001001010.11e-2- Dose-ResponseData legend Generated with Quest Graph™ Renilla Luciferase (pg/well/100µL) RLU Hover mouse to interact
Renilla Luciferase was measured with Amplite™ Renilla Luciferase Reporter Gene Assay Kit in a white 96-well plate with a NOVOstar plate reader (BMG Labtech).
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12536 $950


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Additional Ordering Information
Telephone: 1-800-990-8053
Fax: 1-408-733-1304
Email: sales@aatbio.com
International: See distributors





Overview

Ex/Em (nm)429/466
Storage Freeze (<-15 °C)
Minimize light exposure
InstrumentsLuminescence microplate reader
Category Cell Biology
Reporter Gene Enzymes
Related Tag Enzymes
Common reporter genes include beta-galactosidase, beta-glucuronidase and luciferase. The most versatile reporter gene is the firefly luciferase. Recently there is steadily increasing use of other luciferases, such as Renilla luciferase since these reporters are smaller and do not require the presence of ATP. Our Amplite™ Renilla Luciferase Reporter Gene Assay Kit is designed to provide a fast and sensitive method to detect the luciferase from sea pansy (Renilla reniformis). It uses a proprietary luminogenic formulation to quantify Renilla luciferase activity in cell-based assays. Our formulation generates a luminescent product that gives strong luminescence upon interaction with Renilla luciferase. The kit provides all the essential components. It has high sensitivity and can be performed in a convenient 96-well and 384-well microtiter-plate format. The "glow-type" signal with a half-life of one hour provides a consistent signal across large number of assay plates. The assay is compatible with standard cell growth media. This kit enables the measurement of primary expression or gene expression with wild type and the synthetic hRluc genes .




Protocol


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This protocol only provides a guideline, and should be modified according to your specific needs.
At a glance

Protocol summary

  1. Prepare cell plates
  2. Treat cells as needed
  3. Remove medium from cell plates
  4. Add Renilla Luciferase working solution (100 µL/well for 96-well plate or 25 µL/well for 384-well plate)
  5. Incubate at room temperature for 5 - 10 minutes
  6. Monitor luminescence intensity

Important notes
Thaw all the kit components to room temperature before use. For all luminescent experiments, it is recommended to use white plates to get the best results.

Key parameters
Instrument:Luminescence microplate reader
Recommended plate:Solid white
Preparation of working solution

1. Add one volume of 100X Luciferase Substrate (Component A) to 100 volumes of Assay Buffer (Component B) to make Renilla Luciferase working solution. Note: The reconstituted Renilla Luciferase working solution is very sensitive to light, should be kept from light. In addition, it is not stable, should be prepared fresh, kept on ice and used within 2 hours.


For guidelines on cell sample preparation, please visit
https://www.aatbio.com/resources/guides/cell-sample-preparation.html

Sample experimental protocol
  1. Treat cells (or samples) with test compounds by adding 10 µL of 10X test compounds (96-well plate) or 5 µL of 5X test compounds (384-well plate) in desired compound buffer.
     
  2. Incubate the cell plate in a 5% CO2 incubator at 37°C for desired period of time, typically 4 hours to overnight.

  3. Remove the medium completely.

  4. Add 100 µL (96-well plate) or 25 µL (384-well plate) per well of Renilla Luciferase working solution.

  5. Incubate the plate at room temperature for 5 - 10 minutes. Protect from light.

  6. Monitor luminescence intensity with a luminometer.

 

Example data analysis and figures

The reading (RLU) obtained from the blank standard well is used as a negative control. Subtract this value from the other standards' readings to obtain the base-line corrected values. Then, plot the standards' readings to obtain a standard curve and equation. This equation can be used to calculate Renilla Luciferase samples. We recommend using the Online Linear Regression Calculator which can be found at:

https://www.aatbio.com/tools/linear-logarithmic-semi-log-regression-online-calculator

Figure 1. Renilla Luciferase was measured with Amplite™ Renilla Luciferase Reporter Gene Assay Kit in a white 96-well plate with a NOVOstar plate reader (BMG Labtech).

Disclaimer
AAT Bioquest provides high-quality reagents and materials for research use only. For proper handling of potentially hazardous chemicals, please consult the Safety Data Sheet (SDS) provided for the product. Chemical analysis and/or reverse engineering of any kit or its components is strictly prohibited without written permission from AAT Bioquest. Please call 408-733-1055 or email info@aatbio.com if you have any questions.





References & Citations

Reassembly of a bioluminescent protein Renilla luciferase directed through DNA hybridization
Authors: Cissell KA, Rahimi Y, Shrestha S, Deo SK.
Journal: Bioconjug Chem (2009): 15

RNA detection using peptide-inserted Renilla luciferase
Authors: Andou T, Endoh T, Mie M, Kobatake E.
Journal: Anal Bioanal Chem (2009): 661

The cAMP-dependent protein kinase inhibitor H-89 attenuates the bioluminescence signal produced by Renilla Luciferase
Authors: Herbst KJ, Allen MD, Zhang J.
Journal: PLoS One (2009): e5642

Bioluminescent indicators for Ca2+ based on split Renilla luciferase complementation in living cells
Authors: Kaihara A, Umezawa Y, Furukawa T.
Journal: Anal Sci (2008): 1405

Coelenterazine-binding protein of Renilla muelleri: cDNA cloning, overexpression, and characterization as a substrate of luciferase
Authors: Titushin MS, Markova SV, Frank LA, Malikova NP, Stepanyuk GA, Lee J, Vysotski ES.
Journal: Photochem Photobiol Sci (2008): 189

Mutational optimization of the coelenterazine-dependent luciferase from Renilla
Authors: Woo J, von Arnim AG.
Journal: Plant Methods (2008): 23

Structure-function studies on the active site of the coelenterazine-dependent luciferase from Renilla
Authors: Woo J, Howell MH, von Arnim AG.
Journal: Protein Sci (2008): 725

Crystal structures of the luciferase and green fluorescent protein from Renilla reniformis
Authors: Loening AM, Fenn TD, Gambhir SS.
Journal: J Mol Biol (2007): 1017

Hormone treatment enhances WT1 activation of Renilla luciferase constructs in LNCaP cells
Authors: Hanson J, Reese J, Gorman J, Cash J, Fraizer G.
Journal: Front Biosci (2007): 1387

Quantification of dynamic protein complexes using Renilla luciferase fragment complementation applied to protein kinase A activities in vivo
Authors: Stefan E, Aquin S, Berger N, Landry CR, Nyfeler B, Bouvier M, Michnick SW.
Journal: Proc Natl Acad Sci U S A (2007): 16916


View More Citations




Additional Documents

 
Safety Data Sheet (SDS)


Catalogs
1. Enzyme Probes & Assay Kits

Application Notes
1. AssayWise Letters 2013, Vol 2(1)

Certificate of Analysis