Amino acid (aa) transport through the blood brain barrier (BBB) is relevant both for maintaining brain aa homeostasis and for the distribution of drugs and diagnostic markers such as tracers used for positron emission tomography (PET). Entry and efflux of aas is mediated by specific transporters expressed at the luminal and/or abluminal membranes of brain microvascular endothelial cells (MVECs). To address the mRNA expression of BBB aa transporters, microarray data from three human sources of brain MVECs were compared. This approach confirmed the previously described expression of the neutral aa exchanger Slc7a5-Slc3a2 among other transporters. Based on these results, we are examining the protein expression and membrane localization of transporters in mouse brain tissue sections using immunofluorescence. Further, we are investigating the transport of a new PET tracer, O-(2-[18F]fluoroethyl)-D/L-tyrosine (D/L- FET) by assaying the inhibition of substrate uptake by selected transporters exogenously expressed in Xenopus oocytes. Our results show that both FET isomers inhibit the uptake of substrate by Slc16a2 and Slc36a1, whereas only the L-FET inhibits uptake by Slc7a5-Slc3a2 and Slc7a8-Slc3a2. In summary, we are characterising the expression and function of BBB aa transporters using a strategy of mRNA analysis combined with testing protein expression, localization and function.