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Firefly luciferase

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Firefly luciferase is a bioluminescent enzyme extracted from the firefly. It catalyzes the oxidation of luciferin, a substrate, resulting in the emission of light. The core function of firefly luciferase is to serve as a reporter or marker in various biological and biochemical applications.

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Lab products found in correlation

D luciferin, by Thermo Fisher Scientific (3 mentions) Iblot, by Thermo Fisher Scientific (1 mentions) Mecp2, by Cell Signaling Technology (1 mentions) Pcamkii t286, by Cell Signaling Technology (1 mentions) Dialysis cassette, by Thermo Fisher Scientific (1 mentions) Luciferin, by Thermo Fisher Scientific (1 mentions) Glowmax, by Promega (1 mentions) Dual luciferase assay kit, by Promega (1 mentions)

Close spelling variants of this product

Pierce Firefly Luciferase Flash Assay Kit PMIR-REPORT firefly luciferase miRNA expression reporter vector PMIR-REPORT Firefly Luciferase reporter vector Firefly Luciferase Bioluminescence Assay PMIR-REPORT firefly luciferase vector Pierce Firefly Luciferase Glow Assay Renilla-Firefly Dual-Luciferase Assay kit Firefly luciferase-based ATP determination kit Pierce™ Cypridina-Firefly Luciferase Dual Assay Kit PMIR Firefly luciferase reporters

6 protocols using firefly luciferase

1

Intravascular ATP Release Imaging

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Optical imaging of luciferin-luciferase activity was performed to temporally and spatially assess intravascular ATP release. Mice were injected intraperitoneally with 3 mg D-luciferin (Thermo Fisher, Waltham, Massachusetts) by intraperitoneal route immediately before therapeutic US with MB cavitation. Firefly luciferase (25 μg, Thermo Fisher) was injected by intravenous route 1 min prior to completion of the 10 min US exposure. Optical imaging (IVIS Spectrum, Caliper Life Sciences Hopkinton, Massachusetts) was performed at 5, 10, 15, and 20 min after completion of US using medium binning. Data were expressed as photons/s/cm2.
Mason O.R., Davidson B.P., Sheeran P., Muller M., Hodovan J.M., Sutton J., Powers J, & Lindner J.R. (2019). Augmentation of Tissue Perfusion in Patients With Peripheral Artery Disease Using Microbubble Cavitation. JACC. Cardiovascular imaging, 13(3), 641-651.
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2

Quantification of Signaling Proteins

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Western blots were performed using antibodies against NFAT1(Cell
Signalling, 5861S), pS6K(T389) (Cell Signaling, 9234S), pCamKII(T286) (Cell
Signalling, 12716S), MeCP2 (Cell Signalling, 3456S), pMeCP2(S421)(Rockland,
600-401-X14), Firefly luciferase (Thermo, PA5-32209). The Electrophoresis
and Membrane-Transfer were performed by iBlot (Invitrogen). Imaging was
obtained by Odyssey (Li-Cor) system, following the manufacturer’s
protocol.
Wang H., Tian L., Liu J., Goldstein A., Bado I., Zhang W., Arenkiel B.R., Li Z., Yang M., Du S., Zhao H., Rowley D.R., Wong S.T., Gugala Z, & Zhang X.H. (2018). The osteogenic niche is a calcium reservoir of bone micrometastases and confers unexpected therapeutic vulnerability. Cancer cell, 34(5), 823-839.e7.
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3

Immunofluorescent Microscopy of Mouse Brain and Pancreas

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Thin sections (5–10 µm) from mouse brain or pancreas were obtained and analyzed by immunofluorescent microscopy, as described previously (26 (link)). Every fifth section was analyzed with anti–β-galactosidase (β-gal) for the PRV-BaBlu map. PRV-Ba2001 was analyzed by anti-green fluorescent protein (GFP). Stereotaxic injections were verified by anti-fLuc or anti-HKI. Secondary antibodies were from Jackson ImmunoResearch (West Grove, PA). The primary antibodies used were as follows: β-gal and GFP (Abcam, Cambridge, MA); Cre-recombinase (Novagen/Millipore, Billerica, MA); insulin (Linco/Millipore, Billerica, MA); glucokinase (GK) (Santa Cruz Biotechnology, Dallas, TX); firefly luciferase (Thermo Fisher Scientific, Bannockburn, IL), and hexokinase-1 (HK1) (Cell Signaling, Danvers, MA). For all primary antibodies, immunofluorescent analysis with controls was conducted on adjacent brain sections for autofluorescence, secondary antibody alone, and nonimmune antibody preparations for the appropriate species. Anatomical brain areas were defined by the atlas of Franklin and Paxinos (27 ) using DAPI-positive nuclei and hematoxylin-eosin staining from adjacent sections. An average of at least 140 brain sections/animal were assessed from four or more animals.
Rosario W., Singh I., Wautlet A., Patterson C., Flak J., Becker T.C., Ali A., Tamarina N., Philipson L.H., Enquist L.W., Myers MG J.r, & Rhodes C.J. (2016). The Brain–to–Pancreatic Islet Neuronal Map Reveals Differential Glucose Regulation From Distinct Hypothalamic Regions. Diabetes, 65(9), 2711-2723.
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4

Optical Imaging of Muscle ATP Production

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In 6 mice, optical imaging with a luciferin-luciferase activity was performed to quantify regional ATP production after catheter-based US exposure of the proximal hindlimb muscles (Belcik, et al. 2017 (link)). Mice were injected with 3 mg D-luciferin (Thermo Fisher, Waltham, MA) by intraperitoneal route immediately before US; and firefly luciferase (5 μg, Thermo Fisher) was injected by IV route 1 min prior to completion of US exposure. Optical imaging (IVIS Spectrum, Caliper Life Sciences) was performed 5 minutes after completion of US using medium binning and data were expressed as photons/s/cm2. Release of ATP from erythrocytes, which has been described during high power US cavitation of microbubbles (Belcik, et al. 2017 (link)), was also assessed. Dialysis cassettes (Thermo Fisher) were filled with 3 mL of heparinized blood from normal human volunteers. Cassettes were exposed to US by positioning the EkoSonic catheter adjacent to the membrane using US coupling gel. A total of 4 experiments were averaged and, for each, a control experiment without US was performed. After 5 minutes of US, optical imaging was performed immediately after adding luciferase (10 μg) and D-luciferin (1.2 mg) to each cassette.
Muller M.A., Xie A., Qi Y., Zhao Y., Ozawa K., Noble-Vranish M, & Lindner J.R. (2020). REGIONAL AND CONDUCTED VASCULAR EFFECTS OF ENDOVASCULAR ULTRASOUND CATHETERS. Ultrasound in medicine & biology, 46(9), 2361-2369.
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5

Intravascular ATP Release Imaging

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Optical imaging of luciferin-luciferase activity was performed to temporally and spatially assess intravascular ATP release. Mice were injected IP with 3 mg D-luciferin (Thermo Fisher, Waltham, MA) by IP route immediately before therapeutic US with MB cavitation. Firefly luciferase (25 μg, Thermo Fisher) was injected by IV route 1 min prior to completion of the 10 min US exposure. Optical imaging (IVIS Spectrum, Caliper Life Sciences) was performed at 5, 10, 15, and 20 min after completion of US using medium binning. Repeat injections of D-luciferin and luciferase were repeated 24 hrs later. Data were expressed as photons/s/cm2.
Belcik J.T., Davidson B.P., Xie A., Wu M.D., Yadava M., Qi Y., Liang S., Chon C.R., Ammi A.Y., Field J., Harmann L., Chilian W.M., Linden J, & Lindner J.R. (2017). AUGMENTATION OF MUSCLE BLOOD FLOW BY ULTRASOUND CAVITATION IS MEDIATED BY ATP AND PURINERGIC SIGNALING. Circulation, 135(13), 1240-1252.
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6

Bioluminescent ATP Quantification

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Homogenized tissue was lysed in a lysis buffer (Promega Dual Luciferase Assay Kit) and briefly centrifuged to assess ATP content in target tissues. The supernatant was mixed with a reaction mix containing Firefly Luciferase and Luciferin (Thermo Scientific). The measured luminescence is a direct result of the presence of free ATP, which fuels the conversion of Luciferin to OxyLuciferin by the Luciferase. Luminescence was determined in the Promega GlowMAX.
Hauffe R., Rath M., Schell M., Ritter K., Kappert K., Deubel S., Ott C., Jähnert M., Jonas W., Schürmann A, & Kleinridders A. (2021). HSP60 reduction protects against diet-induced obesity by modulating energy metabolism in adipose tissue. Molecular Metabolism, 53, 101276.
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