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The ONE-Glo™ + Tox Assay combines luciferase assay chemistry with a cell viability marker to better understand reporter gene expression in the context of cell health. The assay uses a two-step, addition-only process to make these measurements in a single well of a plate, negating the need to run parallel assays.
The first part of the assay is a nonlytic fluorescence assay (CellTiter-Fluor™ Cell Viability Assay) that measures the relative number of live cells in a culture population after experimental manipulation. The CellTiter-Fluor™ Assay measures a conserved and constitutive protease activity wit...
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The first part of the assay is a nonlytic fluorescence assay (CellTiter-Fluor™ Cell Viability Assay) that measures the relative number of live cells in a culture population after experimental manipulation. The CellTiter-Fluor™ Assay measures a conserved and constitutive protease activity within live cells and therefore serves as a marker of cell viability. The live-cell protease activity is restricted to intact viable cells and is measured using a fluorogenic, cell-permeant peptide substrate (glycylphenylalanyl-aminofluorocoumarin; GF-AFC). The substrate enters intact cells where it is cleaved by the live-cell protease to generate a fluorescent signal proportional to the number of living cells. This live-cell protease becomes inactive upon loss of cell membrane integrity and leakage into the surrounding culture medium. Fluorescence of the free AFC fluorophore is measured with a microplate reader or CCD imager using an excitation wavelength of 380–400nm and emission wavelength of 505nm.
The second part of the assay uses the ONE-Glo™ Luciferase Assay System to quantify firefly luciferase reporter gene expression from cells made to express this reporter enzyme. The ONE-Glo™ Luciferase Assay Buffer and ONE-Glo™ Luciferase Assay Substrate, provided with this system, are combined to form the ONE-Glo™ Reagent. Ideally suited for high- and ultrahigh-throughput applications, the ONE-Glo™ Assay contains a new fluoroluciferin substrate, resulting in a reagent that is more stable, more tolerant to sample components, and has less odor than standard luciferase assay reagents. Luminescence is measured with a microplate reader or CCD imager.
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ONE-Glo™ + Tox Luciferase Reporter and Cell Viability Assay
ONE-Glo™ Luciferase Assay Buffer
ONE-Glo™ Luciferase Assay Substrate
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This product is available through the Promega Helix onsite stocking program in a –20°C Helix Freezer. The program offers numerous convenient solutions to meet your lab's needs. Helix Freezers are available in two sizes: 5.7 cubic ft. or 9.7 cubic ft.
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Store the ONE-Glo™ + Tox Luciferase Reporter and Cell Viability Assay components at –20°C. Please refer to the Technical Manual for other storage options.
For product intended use please see Patents & Disclaimers tab.
1 × 104 (96-well plate; Panel A) or 5 × 103 (384-well plate; Panel B) cells expressing NFAT response element were treated with serial titrations of ionomycin in the presence of PMA for 6 hours following the example protocols in Section 4.B and 4.C of the Technical Manual. At specific concentrations, ionomycin and PMA work cooperatively to stimulate NFAT-dependent gene expression. However, higher concentrations of ionomycin result in cytotoxicity seen as a decrease in viability (fluorescence). A decrease in reporter expression (luciferase) activity is also observed due to the increase in cytotoxicity.
E7110, E7120 For Research Use Only. Not for Use in Diagnostic Procedures.
E7110, E7120 U.S. Pat. Nos. 7,416,854, 7,553,632 and other patents pending.
E7110, E7120 Patent Pending.
E7110, E7120 Certain applications of this product may require licenses from others.
E7110, E7120 The method of recombinant expression of Coleoptera luciferase is covered by U.S. Pat. Nos. 5,583,024, 5,674,713 and 5,700,673.
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