To assess LUC-2 firefly luciferase activity, treated cells were washed with PBS and lysed with 9 mM MgCl2 and 0.125% Nonidet P-40 substitute in PBS for 15 min at room temperature with shaking. Using half of the lysate, luciferin (PerkinElmer, Waltham, MA) was added to 150 μg/mL for 10 min before the luminescence was read. For Renilla luciferase activity, cells were lysed and luminescence was assessed by using the Renilla Luciferase Glow Assay Kit (Thermo Fisher Scientific, Waltham, MA) and the manufacturer’s protocol.
Renilla luciferase glow assay kit
Manufactured by Thermo Fisher Scientific
The Renilla Luciferase Glow Assay Kit is a laboratory tool used to measure the activity of the Renilla luciferase reporter gene. It provides a bioluminescent signal that can be detected using a luminometer. The kit includes the necessary reagents to perform the assay.
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3 protocols using renilla luciferase glow assay kit
1
Firefly and Renilla Luciferase Assay
2
Hypoxia Response Element Assay
3
Hypoxia Reporter Assay with Exosomes
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The eLUCidate HRE reporter cell line, a stably transfected HeLa cell line which expresses Renilla luciferase reporter gene under the transcriptional control of the hypoxia response element (HRE), was obtained from Genlantis (San Diego, CA). The cells were seeded in a 96well plate at 1x10 4 cells per well and treated with various exosomes at 50 μg/ml for 48 hours. The HRE signals were detected by using a Renilla Luciferase Glow Assay Kit (Thermo Fisher Scientific). Briefly, the cells were rinsed with PBS once. Cell lysis buffer was added at 50 μl/well followed by shaking on a shaker for 15 min. For each well, 20 μl of the cell lysate was transferred to an opaque 96-well plate and incubated with 50 μl of working solution for 10 min. The light output was detected with a SpectraMax M5 microplate reader (Molecular Devices, San Jose, CA).
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