MDR1-MDCK cells were seeded into 24 well Transwell plates and cultured for 3 days to form monolayers. The test compounds were prepared at 10 µM in Hanks' Balanced Salt Solution containing 25 mM HEPES and loaded into the donor compartments of Transwell plates bearing the cell monolayers (pH 7.4 for both donor and receiver compartments). Lucifer Yellow was added to the apical buffer in all wells to assess integrity of the cell monolayer. Duplicate wells were prepared and incubated at 37°C in a CO2 incubator. Samples were removed at time zero and 60 minutes and test compound analysed by LC-MS/MS. Concentrations of Lucifer Yellow in the samples were measured using a fluorescence plate reader. The apparent permeability (Papp) values of test compound were determined for both the apical to basal (A>B) and basal to apical (B>A) permeation and the efflux ratio (B>A: A>B) determined.