Luciferase (Photinus pyralis, Firefly)

Analysis of gene expression is most commonly assayed by transient transfection. These systems are usually based on the use of fusion genes which are inserted into cells, and expression of the gene is assayed within 48 hours after introduction of DNA. Usually the fusion consists of the promoter binding site or enhancer sequence under study which is attached to a reporter gene. The amount of the reporter protein synthesized under the experimental conditions, is presumed to reflect the ability of the sequences studied to direct or promote transcription. Several enzymes are commonly used as reporter proteins, among them are chloramphenicol acetyl transferase (CAT), â-galactosidase, human growth hormone (hGH), and luciferase. Luciferase has become one of the more widely used reporter enzymes. The reporter plasmid contains the gene from the firefly Photinus pyralis. The enzyme catalyzes a bioluminescent reaction which requires the substrate luciferin as well as Mg+2 and ATP. Mixing these reagents with the cell extract containing luciferase, results in a flash of light that decays rapidly. This light can be detected by a luminometer. The total light emission is proportional to the luciferase activity of the sample. The luciferase assay is fast and sensitive and does not require a radioactive substrate as is in the CAT assay. A disadvantage of the luciferase assay is that it requires a rather expensive instrument, the luminometer, to measure enzyme activity. In addition, this assay lacks reproducibility between samples, largely due to the rapid kinetics of the emission.\n\nCellular Localization: Peroxisomal\n\nApplications: \nSuitable for use in Immunofluorescence. Other applications not tested.\n\nRecommended Dilution:\nOptimal dilutions to be determined by the researcher.\n\nStorage and Stability:\nMay be stored at 4 degrees C for short-term only. For long-term storage and to avoid repeated freezing and thawing, aliquot Store at -20 degrees C. Aliquots are stable for at least 12 months at -20 degrees C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.

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SPECIFICATIONS

Catalog Number

L5999-09F

Size

250ul

Applications

IF

Hosts

Rabbit

Form

0.01M PBS. pH 7.4

P Type

Pab

Purity

Purified

Isotype

IgG

References

1. De Wet, J. R., Wood, K.V., et al., Mol. Cell Biol. 7, 725 (1987).

Additional Info

The use of an antibody to detect luciferase can provide an alternative detection assay. This assay directly detects luciferase protein levels, and thus has the advantage that it does not require luciferase activity and is not dependent on rapid kinetics. Moreover, antibodies can detect the luciferase enzyme expression in situ, providing a means to study the localized signal sequences using luciferase as a reporter gene. Species Crossreactivity: Crossreacts with Firefly (Photinus pyralis) Luciferase.