Studies of inhibition of the organic anion transporter used HT-29 cells that were seeded and grown to 70% confluency on 35 x 10 mm clear wall glass bottom dishes (Pelco, USA). Inhibition of the organic anion transporter was achieved by pre-incubating cells with 100 μM sulfobromopthalein (BSP) in media for 1 hour at 37 °C and 5% CO2 in a humidified incubator. A control batch of cells was not treated with BSP. After two washes with 1X PBS, both batches of cells were incubated with 5 μM ICB in media for 2 hours at 37 °C and 5% CO2 in a humidified incubator. Next, the cells were washed twice with 1XPBS and fixed with 4% cold paraformaldehyde for 20 minutes at room temperature. After a single wash with 1XPBS, the cells were co-stained with 3 μM Hoechst 33342 for 10 minutes. After a single wash of 1XPBS, the cells were imaged using a Keyence BZ-X810 fluorescence microscope using a UV filter (ex: 387/11, em: 447/60) and Cy5 filter (ex: 604/40, em: 672/50). For each micrograph, a background subtraction with a rolling ball radius of 100 pixels was applied using ImageJ2 software. After calculating the average MPI for each micrograph, GraphPad Prism was used to plot the data. For each condition, a total of six micrographs were analyzed. The experiments were conducted in triplicate.