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Firefly Luciferase Assay reagent (FLAR-1)
Please call for special pricing on bulk reagent purchase. Product Description:The luciferase assay kit FLAR-1 is a homogenous luciferase gene detection system. Targeting Systems has developed this FLAR-1 luciferase assay kit to facilitate measurement of luciferase activity in HTS (high throughput applications). The kit supplied by Targeting Systems are one step assay kit, i.e. the reagent has all the ingredients necessary for lysing cells as well as the luciferin substrate and stabilizers of the luciferase reaction in a single solution unlike other kits where the assay buffer has to be mixed with the luciferin substrate provided in a separate bottle. The FLAR-1 luciferase assay kit can be used to measure luciferase activity in pre-lysed cell extracts or it can be added directly to the cells (see attached protocol sheet). The assay is compatible with both sample processing robots and with reagent injectors in many luminometers Note: The FLAR-1 reagent can be used to measure firely luciferase activity in both supernatants or lyses of cells transfected with expression vectors expressing firefly luciferase either as an intracellular or secreted form. Principle of the Luciferase AssayThe luciferase assay kit is based upon the bioluminescent measurement of firefly luciferase. This enzyme catalysis the formation of light from ATP and luciferin according to the following reaction:  The intensity of light emission is linearly related to the amount of luciferase and is measured using a luminometer. Luciferase is the most widely used genetic reporter in studies on gene expression due to it’s high sensitivity dynamic range and its natural absence from mammalian cells. The luciferase assay kit offered by Targeting Systems offers the advantages of high sensitivity, consistent reproducibility and cost effectiveness along with the added convenience of a one step assay. The FLAR-1 luciferase assay reagent has been formulated with the cell lysis components, the luciferrin substrate and the bioluminescence stabilizers all included in a single assay reagent. Specifications:Linear range- Assay linear over seven orders of magnitude Limit of detection – less than 1 fg of luciferase per sample Luciferase assay kit (FLAR-1): This kit can be conveniently used for measuring luciferase activity in 96-well plates (or in preparations of lysed cells). As seen in the figure below, the bioluminescent signal is quite stable with a half life greater than 60 mins. The FLAR-1 luciferase assay kit can also be used to measure luciferase activity in prelysed cell extracts. In this case 5-50 µl of the cell lysate is mixed with 100 ul of the luciferase FLAR-1 assay reagent, mixed well and read immediately in the luminometer. The FLAR-1 reagent includes several stabilizers of the luciferase enzyme in its composition. Since the stabilizers used are not luciferase inhibitors a stable bioluminescent signal  Figure 1: Stability of the bioluminescent signal using the FLAR luciferase assay reagent). Results are expresses as the mean of quadruplicate determinations (CV less than 5%). The intensity of the bioluminescent signal using the FLAR reagent from Targeting Systems is comparable to the intensity of the bioluminescent signal in assays using Promega's Bright glo reagent. In the experiment shown above the FLAR reagent was directly added to the supernatant cell culture media Protocol: ASSAY IN 96-WELL FORMAT: Add 100 µl of the FLAR-1 reagent to 50 µl or 100 µl of the supernatant culture media from cells grown in multi-well tissue culture dishes. Incubate for at least 10 minutes and read in a lumionometer or micro plate reader. ASSAY OF LUCIFERASE ACTIVITY IN CELL LYSATES: The FLAR-1 luciferase assay reagents can also be used to measure luciferase activity in pre-lysed cells. Mix 5-50 µl of luciferase containing sample or cell lysate with 100 µl of the luciferase assay kit (FLAR-1) and read immediately in the luminometer. All assay reagents should be close to room temperature at the time of assay. NOTE: If you need to lyse cells first use the cell lysis buffer from Targeting Systems or the Cell culture lysis reagent from Promega (E1531). Intracellular Firefly luciferase activity Lyse cells using our lysis buffer (Catalog no 5X CLR-01). Follow cell lysis protocol supplied with the product. Assay as above using 5 ul to 10 ul of lysate NOTE: If you need to measure intracellular luciferase activity, lyse cells first using the cell-lysis buffer from Targeting Systems. (catalog no 5X CLR-01)
Custom Reagents:Please check out our website www.targetingsystems.net for novel luciferase – based multiplexed assays which enable analysis of up to four promoter activities in the same group of transfected cells. |