What Is A Luciferase Assay
Luciferase reporter assays provide researchers with increased sensitivity and precision due to low signal-to-noise ratios making it easier for researchers to properly process their data and render it meaningful.
Bioluminescence technologies such as firefly luciferase reporter assays provide researchers with increased sensitivity and precision compared to fluorescence assays. Bioluminescence is a chemical process in living organisms in which an enzyme breaks down a substrate to produce light. Unlike with fluorescence, there is no inherent background luminescence in mammalian cells, which allows a luciferase assay to be extremely sensitive with a greater dynamic linear-response range.
Attributes of Luciferase Reporter Assay Technology
INDIGO’s reporter assays utilize bio-luminescence as the detection method. Because there is no inherent background luminescence in mammalian cells or culture media, the sensitivity and dynamic linear-response range of luminescence assays are superior to those of analogous fluorescence-based assays.
Of particular importance to HTS users, it is very rare to encounter a test compound with inherent light emission properties (i.e., chemi-luminescence or phosphorescence). Hence, luminescence-based assays do not suffer from false signals emanating from the test compounds themselves. This is in sharp contrast to fluorescence-based assays.
It is not uncommon for test compounds, regardless of their derivation (such as synthetic small molecules or natural products), to exhibit some level of endogenous fluorescence. Colored test compounds can be particularly problematic. Further, when deployed in primary screening campaigns, test compounds are typically assayed at concentrations 1,000 to 10,000-fold higher than the physiological concentration of natural ligands. Therefore, test compounds producing even low-level fluorescence may contribute inordinately high background, resulting in an erroneous quantitative score (false-positive or false-negative) in a functional assay.