Figure 7.

Bioflavonoid effects on assembly. A. Huh-7.5 cells were infected immediately treated with 0.1μg/ml of Brefeldin A (BFA). Thirty-one hours later, cells were treated with 125μM bioflavonoid for 5 hours after which, the medium was removed and saved for further analysis, and cells were washed twice with PBS to remove bioflavonoids. Fresh medium was added, and cells were subjected to three cycles of freeze/thaw. After clearing cellular debris, supernatants were used to infect naïve cells followed by luciferase assays 72 hours later. All values were normalized to ‘DMSO + BFA’. B. Control experiment to determine if bioflavonoids effect viral protein translation in the conditions of part A. Huh-7.5 cells were infected and treated with BFA and bioflavonoids exactly as described in part A. Immediately after the 5-hour bioflavonoid treatment, cells were washed and lysed, and Renilla luciferase assay was performed. C. Control experiment to determine the effect of BFA on viral secretion in the original part A assembly assay. The supernatants saved from the original culture were concentrated 30-fold to remove bioflavonoids and used to infect naïve cells, followed by luciferase assays 72 hours later. All values were normalized to DMSO treatment. (*, **, and *** indicate P<0.05, P<0.005, and P<0.0005, respectively. Error bars reflect standard deviation.)