Luciferase Reporter Gene Assay Kit

Catalog No. ABIN1000342

Product Details

Application: Luciferase Reporter Gene Assay (LRA)

Characteristics: High sensitivity and wide detection range: detection of as little of 2 fg luciferase and as few as 4 cells. Plus, the emitted light is linear over seven orders of magnitude.
Compatible with routine laboratory and HTS formats: assays can be performed in tubes or microplates, on LJL Analyst, Berthold Luminometer, Top-Count, MicroBeta counters, chemiluminescent image plate readers (CLIPR/LeadSeeker). Assay reagents compatible with all liquid handling systems.
Fast and convenient: homogeneous mix-and-measure assay allows detection of luciferase levels within 10 minutes. The optimally combined reagent system allows a single addition step, and simultaneous cell lysis and detection.
Robust and amenable to HTS: Z' factors of 0.6 to 0.8 are observed in 96- well and 384-well plates. Can be readily automated on HTS liquid handling systems.

Application Details

Application Notes: Gene Regulation: gene expression level, characterization of promoter activity, modulation of gene expression by receptors, transcription factors and small molecules.
Drug Discovery: high-throughput screen for gene modulators.

Comment:

Incubation time. Both the luciferin/luciferase reaction and cell lysis are fast, so incubation for 2 to 10 minutes following reagent addition is generally enough for mammalian cells (e.g. HEK293, CHO).
Cell number. The optimized reporter gene assay reagent is very sensitive to luciferase (detection limit 2 fg) and exhibits linearity over seven orders of magnitude. As few as 4 cells can be determined and a linear response is still observed with as many as 80,000 cells per 96- well. For assay optimization, it is recommended that the optimal number of cells per well be determined by serial dilution of cells. Cells can be adherent or in suspension cultures. Cell lysis and mixing. For the sake of convenience, the addition of 1 volume of reconstituted reagent to 1 volume of cells allows a sufficient mixing. No additional mixing is required since the specially formulated buffer instantly lyses mammalian cells.

Protocol: The Luciferase Reporter Gene Assay is based on the bioluminescence generated during the luciferin/luciferase reaction. The reconstituted reagent has been optimized to combine cell lysis and detection into one single step. Phenol red in culture media does not interfere in this assay. All data in the Technical Notes were obtained in media containing phenol red.

Restrictions: For Research Use only

Handling

Storage: -20 °C

References

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Roupelieva, Griffiths, Kremmer, Meisterernst, Viejo-Borbolla, Schulz, Haas: "Kaposi's sarcoma-associated herpesvirus Lana-1 is a major activator of the serum response element and mitogen-activated protein kinase pathways via interactions with the Mediator complex." in: The Journal of general virology, Vol. 91, Issue Pt 5, pp. 1138-49, (2010) (PubMed).

Gentry, Taormina, Pyles, Yeager, Kirtley, Popov, Klimpel, Eaves-Pyles: "Role of primary human alveolar epithelial cells in host defense against Francisella tularensis infection." in: Infection and immunity, Vol. 75, Issue 8, pp. 3969-78, (2007) (PubMed).

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Target Details

Background: Bright bioluminescent reagent system allows rapid quantitation of luciferase reporter gene expression in transfected cells within 2 min.

The Luciferase Reporter Gene Assay is based on the quantitation of luciferase expression in mammalian, yeast or E. coil cells, using luciferin and ATP as substrates. The reaction results in light production which can be conveniently measured on a luminometer. This bioluminescent reporter gene assay is extremely sensitive and is especially suitable for quantifying luciferase expression in recombinant cells. This ultra-sensitive, homogeneous cell-based assay only requires adding a single reagent to the cells and measuring the light intensity after a short incubation step (2 minutes). Assays can be performed in tubes, cuvettes or multi-well plates. All kit components are compatible with culture media and with all liquid handling systems. With an extended luminescence emission kinetics (half-life 40 min), the SuperLight TM luciferase assays are especially suitable for high-throughput screening in 96-well, 384-well and 1536-well plates. In addition, the reagent provided in the kits has been formulated for maximum sensitivity, reproducibility and long shelf-life. Applications for this kit include gene regulation studies and high-throughput screening of gene modulators.