Why You Don’t Need to Select a Wavelength for a Luciferase Assay

Promega kit depicted; test involves wavelength for a luciferase assay.

It’s a question I’m asked probably once a week. “What wavelength do I select on my luminometer when performing a luciferase assay?” The question is a good and not altogether unexpected one, especially for those new to bioluminescent assays. The answer is that in most cases, you don’t and in fact shouldn’t select a wavelength (the exception to this rule is if you’re measuring light emitted in two simultaneous luciferase reactions). To understand why requires a bit of an explanation of absorbance, fluorescence, and luminescence assays, and the differences among them.

Absorbance, fluorescence, and luminescence assays are all means to quantify something of interest, be that a genetic reporter, cell viability, cytotoxicity, apoptosis, or other markers. In principle, they are all similar. For example, a genetic reporter assay is an indicator of gene expression. The promoter of a gene of interest can be cloned upstream of a reporter such as β-galactosidase, GFP, or firefly luciferase. The amount of each of these reporters that is transcribed into mRNA and translated into protein by the cell is indicative of the endogenous expression of the gene of interest.

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The Secret Fluorescent Life of Flying Squirrels

flying squirrel specimen
A flying squirrel museum specimen under normal light versus ultraviolet light. Photo courtesy of AM Kohler, et al.

In May 2017, a surprising discovery was made in the woods of Bayfield County, Wisconsin, just about a 5-hour drive north of Promega headquarters. Jonathan Martin, Associate Professor of Forestry at Northland College, was exploring the forest with an ultraviolet (UV) light in search of fluorescent lichen or plant life. What he found instead was a bright pink glow coming from a most unexpected source—a flying squirrel.

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Surfing the Light Waves: Shrimp, Coral, Turtles and Other Fluorescent Organisms

A branching torch coral, Euphyllia glabrescens.

Have you ever walked on a beach and noticed that the waves seem to glow as they roll onto shore? Perhaps you have seen fish or jellyfish that glow in the dark, or maybe you’ve chased fireflies in your backyard or on a camping trip. These are all forms of luminescence (the production of light without adding heat), but the manner that these organisms produce their light can be quite different.

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