Plasma membrane transporter proteins play an important role in taking up nutrients into and effluxing xenobiotics out of cells to sustain cell survival. In the last decade, a number of studies have shown that these physiologically important transporters affect absorption, distribution, and excretion of major anticancer agents in clinical use. More importantly, many transporters have been reported to be differentially upregulated in cancer cells compared to normal tissues, suggesting that the differential expression of transporters in cancer cells may become good targets for enhancing drug delivery as well as diagnostic markers for cancer therapy. Hence, utilizing the knowledge of transporter functions likely provides us with the possibility of delivering a drug to the target tissues, avoiding distribution to other tissues, and improving oral bioavailability. This chapter focuses on methodology to analyze the activity of transporters that are involved in drug transport.