名称：原装供应| b-Gal Reporter Gene Assay, che
Enzyme reaction: In the first step, the substrate Galacton Plus becomes deglycosylated by the enzymatic activity of the β-galactosidase contained in the sample. This incubation step is performed at pH 7.8, at which bacterial β-galactosidase is highly active.
Specificity: The β-Gal Reporter Gene Assay is designed for specifically measuring bacterial β-galactosidase activity. To achieve this, the enzyme reaction is conducted at a pH that is optimized for bacterial β-Gal, but allows no significant endogenous eukaryotic β-galactosidase activity. However, if very high endogenous β-galactosidase activity is found (e.g., with hepatic cells), a heat treatment can be performed to ensure the specific determination of the amount of bacterial β-galactosidase that is encoded by the transfected plasmid. At this neutral pH, the cleaved dioxetane is protonated and will not produce a light signal.
However, the exact detection limit and measuring range depend on the measuring device and the measuring conditions used. The β-Gal Reporter Gene Assay, chemiluminescent, is used to quantitatively measure β-Gal expression in eukaryotic cells that are transfected with a plasmid that bears the β-Gal-encoding lacZ gene as a reporter. Sample material: Cell extracts. Detection range and sensitivity: Under assay conditions, the detection range for the positive control is between 20 fg and 20 ng (Figure 2). Light reaction: The light reaction is initiated by shifting the pH to a value higher than 12. Due to this shift in pH, the activated intermediate becomes deprotonated and decomposes with the emission of light (475 nm).