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0 Item(s) - $0.00One Step Luciferase Assay System, 10ml
Background:
Luciferase is the general term given to a class of oxidative enzymes that catalyze reactions that give off light, a process known as bioluminescence (Fig. 1). In biology, researchers can take advantage of this reaction and use it as a readout for various biological processes. This has perhaps been exploited most in luciferase reporter cell lines where a promoter region from a gene of interest is placed immediately upstream of the coding sequence for luciferase. In this system, transcriptional activation of the gene of interest leads to a level of luciferase expression that is proportional to the level of gene activation.
The level of activation is then assessed by lysing the cells and adding the luciferase substrate, D-luciferin. Using the One-Step Luciferase Assay System, this is accomplished in a single step as the lysis buffer has been formulated with D-luciferin. After a brief incubation period, bioluminescence is read on a luminometer.
Description:
The One-Step Luciferase Assay System is designed to be used for high-throughput, sensitive quantitation of firefly luciferase activity in mammalian cell culture. The reagent consists of two components, a Luciferase Reagent Buffer (Component A) and Luciferase Reagent Substrate (Component B). Component A and Component B are combined to form a working solution that contains all the necessary components for cell lysis and luciferase quantitation. This assay system has several features:
• Sensitive – highly sensitive detection of firefly luciferase activity.
• Stable – the signal output is stable for more than two hours, providing flexibility with regard to incubation time
• Convenient – simple one-step, homogeneous protocol.
• High-throughput – one-step homogeneous protocol minimizes handling steps to support high-throughput screening applications
• Compatibility – works well with a variety of common media containing 0-10% serum and phenol red.
• Instrumentation – does not require a luminometer with injectors.
• Sensitive – highly sensitive detection of firefly luciferase activity.
• Stable – the signal output is stable for more than two hours, providing flexibility with regard to incubation time
• Convenient – simple one-step, homogeneous protocol.
• High-throughput – one-step homogeneous protocol minimizes handling steps to support high-throughput screening applications
• Compatibility – works well with a variety of common media containing 0-10% serum and phenol red.
• Instrumentation – does not require a luminometer with injectors.
Synonym(s):
Luciferin, Firefly luciferase, luciferase buffer
Format:
COMPONENTS:
Instructions for use:
See product datasheet for detailed protocol.
Storage / Stability:
At least 6 months when stored as directed. Upon first thaw, store in aliquots at -20°C. The reagent may be subjected to several freeze/thaw cycles with no effect on functionality, but it is recommended that freeze/thaw cycles be avoided whenever possible.
Application(s):
• Monitor firefly (Photinus pyralis) luciferase activity in cultured mammalian cells. • High-throughput screening using luciferase reporter cell lines
Notes:
• The reagent has been validated in a 96-well format. Other formats will require scaling and optimization by the end-user.
• Luciferase Reagent Buffer must be at ~ room temperature (20- 25°C) before use.
• Luciferase Assay Reagent should be added to cells for at least 5 minutes before measuring luminescence to allow complete cell lysis.
• For maximal light intensity, measure samples within 1 hour of reagent addition.
• Avoid exposing to excessive heat or light during incubation.
• Different cell lines may exhibit variation in lysis ability and/or luminescence signal and may require slight optimization by the end-user.
• Luminescence signal is affected by assay conditions. Results should be compared between samples measured using the same cell type and media/serum combinations.
• To analyze multiple plates, include a common control sample in each plate and normalize the luminescence of each plate to the control contained in the same plate.
• Background luminescence is a characteristic of luminometer performance, therefore, background luminescence must be subtracted from all readings for accuracy
• Luciferase Reagent Buffer must be at ~ room temperature (20- 25°C) before use.
• Luciferase Assay Reagent should be added to cells for at least 5 minutes before measuring luminescence to allow complete cell lysis.
• For maximal light intensity, measure samples within 1 hour of reagent addition.
• Avoid exposing to excessive heat or light during incubation.
• Different cell lines may exhibit variation in lysis ability and/or luminescence signal and may require slight optimization by the end-user.
• Luminescence signal is affected by assay conditions. Results should be compared between samples measured using the same cell type and media/serum combinations.
• To analyze multiple plates, include a common control sample in each plate and normalize the luminescence of each plate to the control contained in the same plate.
• Background luminescence is a characteristic of luminometer performance, therefore, background luminescence must be subtracted from all readings for accuracy
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