Mono-Lumi Firefly Luciferase Reporter Gene Assay Kit

 

Product Name	Cat. No.	Spec.
Mono-Lumi Firefly Luciferase Reporter Gene Assay Kit	G1702-100T	100T

 

 

The firefly-luciferase system is mostly used for reporter gene detection. The transcriptional regulatory elements of the target gene were cloned upstream and downstream of the luciferase to construct a reporter gene plasmid. The plasmid was then transfected into cells, and after treating the transfected cells with drugs or other stimuli, the cells were collected and lysed, and luciferase activity in the lysate was measured. The transcriptional regulation of target genes by exogenous stimuli such as drugs is determined by luciferase activity.

The Mono-Lumi Firefly Luciferase Reporter Gene Assay Kit has high sensitivity, broad detection range and stable fluorescence signal. Using Firefly Luciferase, which catalyzes the oxidation of firefly luciferin substrate (luciferin) to generate oxyluciferin under the conditions of Mg2+, ATP, O2, etc. and generates biofluorescence in the process, the principle of Firefly Luciferase activity can be rapidly detected.

 

 

Ship with dry ice; store at -80℃ protected from light, valid for at least one year; store at -20℃ protected from light, for up to 6 months.

 

 

Component Number	Component	G1702-100T
G1702-1	Cell Lysis Buffer	50 mL
G1702-2	Luciferase Reaction Substrate (50×)	200 μL
G1702-3	Luciferase Reaction Buffer	10 mL
Manual		One copy

 

 

1. Preparation of 1× firefly luciferase reaction working solution

1.1 Thaw each component of the kit at room temperature and mix upside down to ensure complete dissolution. The Luciferase Reaction Substrate (50×) should be thawed on ice and centrifuged briefly to ensure that the reagent sinks to the bottom of the tube;

1.2 The Luciferase Reaction Substrate (50×) was diluted 50 times with Luciferase Reaction Buffer, i.e., 10 μL Luciferase Reaction Substrate (50×) + 490 μL Luciferase Reaction Buffer was mixed well to obtain 500 μL of 1× Firefly Luciferase Reaction Buffer.

2. Cell pretreatment and lysis

2.1 Plate cells/well in appropriate well plates, perform cell transfection or other relevant pretreatment according to the experimental requirement.

2.2 For adherent cells: After removing the original cell culture medium, add appropriate amount of Cell Lysis Buffer to each well according to the table below.

For suspension cells: After centrifuge, remove the supernatant and add appropriate amount of Cell Lysis Buffer to precipitated cells

Refer to the table below for the amount of lysis buffer suggested .

 

Cell Culture well-Plate	Cell Lysis Buffer/Well
6-well	500 μL
12-well	300 μL
24-well	200 μL
48-well	150 μL
96-well	100 μL

 

2.3 Lysis cells completely at room temperature for 10-20 min, collect cells in a 1.5mL centrifuge tube by centrifuge at 12,000 ×g at 4℃ for 10 min, keep the supernatant (cell lysate) for later use.

3. Firefly luciferase assay

3.1 Pre-prepared 1× firefly luciferase reaction working solution was equilibrated to room temperature and added to an opaque white 96-well plate at 100 μL/well;

3.2 The cell lysis supernatant from step 2 was added to the above 96-well plate at 20 μL/well;

3.3 Shake horizontally to homogeneity and immediately perform a chemiluminescence assay using a Luminometer luminescence meter, a multifunctional enzyme labeler with a chemiluminescence detection module, or other instrument capable of detecting bioluminescence.

 

 

1. Prior to the assay, the pre-prepared 1× Firefly Luciferase Reaction Work Solution needs to be brought back to room temperature before use.

2. Luciferase Reaction Buffer may precipitate after thawing. Please shake thoroughly before use to ensure complete dissolution.

3. The 1× luciferase reaction working solution is prone to be oxidized. For multi-sample detection, the addition of 1× firefly luciferase reaction working solution should be controlled in a short time. It is recommended to add samples with the multichannel pipettes, and keep the volume of each well is consistent.

4. To prevent interference between wells, it is recommended to use a white, opaque well plate. The black, opaque well plate can also be used, but it will absorb the fluorescence signal, reducing the fluorescence signal of detection.

5. For your safty and health, please wear safety glasses, gloves, or protective clothing.

 

For Research Use Only!