Assessment of Multidrug Resistance Associated Protein (MRP1) in RMS cell lines. (A) MRP1 expression determined by indirect immunofluorescense microscopy showing intracellular distribution of MRP1 (green) in RMS cells following treatment with AS-DACA. Co-stained with the nuclear stain DAPI (blue). (B) Levels of MRP1 were determined by Western blot before (-), and after (+), exposure to AS-DACA (2 × IC50 for 16 h). Densitometry was performed on triplicate blots with band intensity normalized for both background noise and total protein loading differences as determined by Ponceau S staining. (C) Influence of (R)-verapamil (1 and 10 μM) on AS-DACA cytotoxicity. RD (top) and Res30 (bottom) cells were treated for 72 h with (R)-verapamil whilst being exposed to serially increasing concentrations of AS-DACA. Error bars in (A) and (C) represent the SEM for three independent experiments.