UA-Glo® Beta-Gal Luminescence Detection Kit

Beta-galactosidase (Beta-Gal) is one of the earliest reporter genes and has a wide range of applications. Traditional methods for detecting Beta-Gal include colorimetric and fluorescence methods, but both have disadvantages such as non-homogeneous detection or low sensitivity. The UA-Glo® Beta-Gal Luminescence Assay Kit uses Lu-Gal as the substrate. After Beta-Gal cleaves Lu-Gal, it generates luciferin, the substrate of luciferase. Luciferin is then acted upon by luciferase to produce luminescence, which is detected. The intensity of the luminescence is directly proportional to the expression level of Beta-Gal. The UA-Glo® Beta-Gal Luminescence Assay Kit is a homogeneous detection reagent. It can be detected as soon as 0.5 hours after adding the reagent, and the signal intensity is very stable, remaining largely unchanged within 3 hours. It is particularly suitable for high-throughput screening of compounds.

The components and specifications of the Beta-Gal luminescence detection kit are as follows：

1. Seed cells expressing Beta-Gal at an appropriate density in white, clear-bottom 96-well or 384-well cell culture plates. Serum, such as FBS, may contain mammalian Beta-Gal, so it is recommended to use heat-inactivated FBS (56°C, 0.5 hr) to prepare the complete medium for cell seeding. Within the same plate, it is advisable to include wells with cells that do not express Beta-Gal or wells with only complete cell culture medium as background controls for Beta-Gal detection. The background readings can be used to correct the experimental readings.

2. Perform cell treatment according to experimental requirements, followed by Beta-Gal detection.

3. Take out the Beta-Gal detection reagent, allow it to completely dissolve, and equilibrate to room temperature (22°C–25°C). Invert gently to mix thoroughly.

4. Remove the cell culture plate and equilibrate it to room temperature (22°C–25°C). The luciferase reaction is sensitive to temperature changes. Both the reagent and test samples/plates should be equilibrated to room temperature, and the temperature should remain constant (±1°C) throughout the testing process.

5. Add Beta-Gal detection reagent equal in volume to the culture medium in each well of the cell culture plate. For example, add 100 µl of detection reagent to 100 µl of culture medium.

6. Shake the plate for 30 seconds to mix thoroughly, then incubate in the dark for 30 minutes before performing luminescence detection. Readings can be taken within 3–5 hours.

7. Data processing: For most experiments, the raw data can be used directly for subsequent calculations. If the background is high, subtract the background readings (from cells without Beta-Gal expression or complete medium-only wells) from the experimental readings, and use the background-corrected data for further calculations.

1. Different batches should not be mixed.
2. Without strict verification, it is not recommended to change the dosage of reaction reagents.
3. For research purposes only.