β-Galactosidase Assay is a method used to measure the activity of the enzyme β-galactosidase, which catalyzes the hydrolysis of β-galactosides into monosaccharides. β-galactosidase is commonly used as a reporter gene in molecular biology to monitor gene expression and cellular activity in various research applications. The typical assay involves incubating a sample containing β-galactosidase with a substrate and then measuring the product concentration, which is directly proportional to the enzyme activity. The assay can be adapted for use in various sample types, including bacterial cultures, cell lysates, or tissue homogenates. The enzyme’s activity is often assessed using synthetic substrates such as ONPG (o-nitrophenyl-β-D-galactopyranoside) or X-Gal (5-bromo-4-chloro-3-indolyl-β-D-galactopyranoside).

The ZELLX® β-Galactosidase Assay Kit to quantify active β-galactosidase levels in cells transfected with plasmids that express the Lac Z gene. Lac Z is a popular reporter gene in transfection experiments due to the robust resistance of β-galactosidase to proteolytic degradation and the simplicity of measuring its activity. This enzyme catalyzes the hydrolysis of ortho-nitrophenyl-β-D-galactopyranoside (ONPG) into ortho-nitrophenol (ONP), which yields a bright yellow color detectable at an absorbance of 420 nm. The intensity of the color produced is directly proportional to the concentration of β-galactosidase, facilitating accurate and straightforward assessments of gene expression.