β-galactosidase, is one of the first and most popular reporter enzymes which are commonly used in cell biology to study transcriptional activity of genes. β-galactosidase, encoded by the lacZ gene of E. coli, catalyzes the hydrolysis of β-galactosides into monosaccharides. It is widely used as a reporter enzyme to study gene expression, protein-protein interactions, and normalization of transfection efficiency.

β-Galactosidase Activity Assay Kit provides a simple, reproducible, and sensitive tool of measuring β-Galactosidase activity in plasma, serum, cell lysates, and other biological liquid samples. In this assay, O-nitrophenyl β-D-galactopyranoside (ONPG) is hydrolyzed into galactose and ortho-nitrophenol in the presence of β-galactosidase. Upon β-galactosidase protease cleavage, the o-nitrophenol produces a yellow color that can be detected at absorbance 420 nm. The optical density measured is directly proportional to the activity of β-Galactosidase present in the sample.

• Microplate reader capable of measuring absorbance at 420 nm
• Incubator at 37°C
• Precision pipettes to deliver 2 μL to 1 mL volumes
• Tubes to prepare sample dilutions
• 15 mL conical tubes

1. Add 20 μL of the diluted β-Galactosidase Standards or samples to the 96-well microtiter plate.
2. Initiate the reaction by adding 70 µL Working Solution to each well. Carefully shake the plate for a few seconds to mix.
3. Incubate the plate for 30 minutes at 37°C with mixing in the dark.
4. Add 120 µL Stop Solution into each well.
5. Measure the absorbance at 420 nm using a plate reader.