Luciferase Reporter Gene Assay, high sensitivity

Uses

The Luciferase Reporter Gene Assay is used to quantitatively measure the expression of firefly luciferase in eukaryotic cells or bacteria transfected with a vector that encodes firefly Photinus pyralis(EC 1.13.12.7) luciferase. The Luciferase Reporter Gene Assay can be used in manual or automated luminometers in a microplate or tube format, as well as in scintillation counters or photographic films. In order to achieve maximum sensitivity, the use of a luminometer that operates with ultra-fast photon counters is recommended (e.g., EG&G Berthold luminometers).

General Description

Principle

The Luciferase Reporter Gene Assay contains all substrate components for the enzymatic determination of firefly luciferase activity, as well as a lysis buffer for the mild and rapid extraction of luciferase from eukaryotic cells. In the first step, transfected cells are lysed by the addition of lysis buffer. Subsequently, the cell extract is transferred to a microplate or tube, depending on the type of equipment used. The enzymatic reaction is then started by adding the mixture of reaction buffer and luciferase substrate (luciferase assay reagent), which contains all of the components required for starting the chemiluminescent reaction. The luciferase assay reagent may be added manually or by automated injector. Photon emission is quantitated using a luminometer or alternative equipment such as a scintillation counter.

The luciferase assay is much more sensitive (100- to 1000-fold) than the standard isotopic CAT (chloramphenicol acetyltransferase) assay. The Luciferase Reporter Gene Assay is not suitable for the determination of luciferases of bacterial origin.

Contents

• Reaction Buffer
• Luciferase Substrate
• Lysis Buffer