Luciferase assays for NF-κB promoter transcriptional activity

HCT116 human colon cancer cells were transfected with the pNL1.3 [secNluc/NF-κB-RE/Puro] reporter plasmid using polyethylenimine MAX MW 40,000 (PolyScience, Warrington, PA, USA). Transfectants stably expressing NF-κB-secNluc were obtained after treatment with puromycin and cloning for use in measuring NF-κB transcriptional activity. The cells were referred to as HCT116-NF-κB-secNluc cells. HCT116-NF-κB-secNluc cells were seeded in 96-well plates (2 × 10⁴ cells/well). After 24 h incubation, the cells were treated with 1, 5 or 10 μM fucoxanthinol for 24 h. Firefly luciferase activity levels were determined using the Nano-Glo® Luciferase Assay System (Promega Corporation). The basal NF-κB luciferase activity in the control was set as 1.0. Data are expressed as the means ± SD (n = 6).