Biluciferase reporter gene kit

The ARE fluorescence activity was tested in cultured BMEC with or without H 2 O 2 via an ARE reporter gene according to the manufacturer's instructions (Promega). In brief, after the treatment of BMEC with H 2 O 2 (600 μM) for 0, 6, 12, and 24 h, 50 μL of luciferase assay reagent II was added to the BMEC, followed by 10 μL of cell lysis buffer. The activity of relative light unit 1 (RLU1) was detected with a biluciferase reporter gene kit (E1910; Promega). Then, 50 μL of 50× Stop and Glo reagent was added to the BMEC, and the activity of relative light unit 2 (RLU2) was detected again with a biluciferase reporter gene kit (E1910; Promega). The RLU1/RLU2 ratio was used to determine the transfection efficiency of ARE.