Fig. 3

Effect of SKN on P-gp activity. a P-gp expression in A2780 and A2780/PTX cells was evaluated using flow cytometry after incubation with FITC-P-gp antibody. b The effect of SKN on P-gp activity was further investigated. Cells (6.0 × 10⁴/well) were treated with different concentration of SKN (0, 0.5, 1, 2 μM), PTX (1 μM) and combination of SKN/PTX (both at 1 μM) for 4 h, then incubated with MDR dye in dark for another 1 h. Verapamil was used as a positive control. Intracellular fluorescence was determined on a SpectraMax M5 microplate reader. c, d The effect of SKN on P-gp efflux activity was investigated. Cells (6.0 × 10⁴/well) were incubated with a specific fluorescent P-gp substrate R123 (0.5 μg/mL) with or without SKN (1 μM) for 1 h, then washed twice with ice-cold PBS and incubated in R123-free medium at 37 °C for additional 1 h with SKN or without SKN. Cells were analyzed on flow cytometry to detect R123 fluorescence. The verapamil (20 μM) was used as the positive controls in a parallel study