Bioluminescent Luciferase Assays

Luciferase is an enzyme that catalyzes a bioluminescent reaction, producing visible light as a byproduct. This enzyme is naturally found in various organisms, including certain insects, fungi, and marine animals, with the firefly being one of the most iconic examples. Notably, luciferase is absent in mammalian cells, making it an excellent reporter enzyme for biological research. Because luciferase activity is highly sensitive, has a wide dynamic range of quantification, and lacks of toxicity, luciferase reporter assays have become a popular reporter enzyme in life science research and drug discovery.

Bioluminescence, Chemiluminescence, and Fluorescence

Chemiluminescence is a chemical process that results in the emission of light as one of products of the reaction. When such a reaction is produced from a biological source like luciferase, it is called bioluminescence. In comparison, fluorescence is the process in which a fluorophore (also known as a fluorescent molecule) such as GFP absorbs light energy at one wavelength to emit light of another wavelength.

Bioluminescence vs Fluorescence. The light from bioluminescence is produced from a chemical reaction between luciferase enzymes and its substrate (for example, firefly luciferase and luciferin). As there are different types of Luciferases that also react differently, cofactors involved may vary. In contrast, in fluorescence, light is generated when a fluorophore absorbs light or radiation (excitation energy) and emits visible response.

Unlike fluorescence, bioluminescence does not require external excitation light, which minimizes background signal and improves sensitivity. This makes bioluminescent luciferase assays more suitable for real-time detection of cellular activity and regulatory events.

Bioluminescence vs Fluorescence in Research

Both bioluminescence and fluorescence are widely used in the laboratory setting. Fluorescent reporters are stable and easy to visualize in living systems but are less suited for dynamic monitoring due to signal persistence. Fluorescence assays can also suffer from cellular autofluorescence, which reduces sensitivity.

Conversely, Bioluminescent reporters offer lower background and higher resolution. For example, here at Signosis, we offer many different TF Specific Reporter Cell Lines, which are designed to report the activity of specific transcription factors (TFs). Because the activity of these TFs can change in real time, bioluminescence would be the preferred reporting system, and these cell lines do that through the use of luciferase.

It is important to know the difference between the many types of luminescence and how they work. Many imaging systems and plate readers today are multi-purpose and are can read multiple types of luminescence, but the filter or excitation settings will differ between assay types. Customers sometimes confuse bioluminescence (which typically does not require excitation or emission filters) with fluorescence (which does). When working with luciferase assays, especially chemiluminescent or firefly luciferase assays, make sure that your reader program does not activate excitation light or select optical filters, as this can interfere with signal detection.

Firefly, Renilla, and Gaussia Luciferase

The most commonly used luciferases, Firefly and Renilla, are typically expressed inside the cell and require lysis to measure bioluminescence. In contrast, a secreted luciferase is naturally released into the cell culture medium, which enables easy detection directly from cell media.

Luciferase	Secreted	Size (kDA)	Substrate base
Firefly	No	61	D-luciferin
Renilla	No	36	Coelenterazine
Gaussia	Yes	20	Coelenterazine

Because Gaussia luciferase is secreted into culture media, it enables repeated, non-invasive measurements of luciferase activity over time. This feature makes secreted Gaussia luciferase cell lines an ideal choice for kinetic assays, time-course studies, and long-term monitoring of biological process. In contrast, firefly luciferase assays typically require cell lysis, offering excellent sensitivity for endpoint assays.

Firefly and Gaussia Luciferase Systems at Signosis

Whether your experiment requires an intracellular firefly luciferase assay for transcriptional analysis or a secreted Gaussia luciferase reporter for real-time kinetic monitoring, Signosis provides optimized, ready-to-use luciferase reporter cell lines to make detection simple and reliable.

Our Firefly Luciferase Reporter Cell Lines are ideal for measuring specific transcription factor activity with strong, quantifiable luminescent signals. Meanwhile, our Secreted Gaussia Luciferase Cell Lines allow continuous sampling without disrupting cell growth.

In addition to luciferase assays and reporter cell lines, Signosis also provides optimized substrates, lysis buffers, and detection reagents designed to ensure high signal-to-noise ratio and compatibility with most plate readers.

From sensitive firefly assays to convenient secreted Gaussia reporters, Signosis provides the tools and reagents you need to advance bioluminescent research. We also offer fully customizable reporter cell lines to meet your specific experimental needs. We also offer customizable luciferase reporter cell lines to meet specific experimental needs. For more information or custom inquiries, please contact us at info@signosisinc.com