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. Author manuscript; available in PMC: 2009 May 1.
Published in final edited form as: Biochem Pharmacol. 2008 Feb 7;75(9):1858–1867. doi: 10.1016/j.bcp.2008.01.017

Figure 2. Reporter Assay of HIF-1 transcriptional activity.

Figure 2

MDA-MB-231 cells grown to 60% confluency in 6-well plates were transfected with the HIF-1 reporter construct, pH3-SVL, using the lipofectamine transfection reagent. Cells were allowed to recover overnight, given the indicated concentrations of DIM and the PHD inhibitors 2-2′dipyridyl (DPL) and dimethyloxaloylglycine (DMOG), and the antioxidants dithiothreitol (DTT) or N-acetyl-cysteine (NAC) and incubated in normoxic or hypoxic conditions for 8 hours. Activity of luciferase in cell lysates was measured by chemiluminescence. Protein concentration in cell lysates was measured using the Bradford assay and results expressed as relative light units (RLU) per mg protein as the average +/− SD for three replicates.