The aim of this study was to explore the potential of a new selective dopamine transporter (DAT) compound as a radioligand for positron emission tomography (PET) examination of DAT in the human brain. The high affinity DAT compound N-(3-iodoprop-2E-enyl)-2β-carbomethoxy-3β-(4-methylphenyl)nortropane (PE2I) was radiolabelled by the O-methylation approach and the binding was characterised by PET in cynomolgus monkeys and a healthy man. Metabolite levels in plasma were measured by gradient high-performance liquid chromatography. O-methylation of the corresponding free acid precursor with [¹¹C]methyl triflate gave high radiochemical yield (80%) and specific radioactivity (55 GBq/μmol). [¹¹C]PE2I binding in cynomolgus monkeys was nine times higher in the striatum than in the cerebellum at peak equilibrium, which appeared 55–65 min after injection. Displacement and pretreatment measurements using unlabelled β-CIT, GBR 12909, cocaine, citalopram and maprotiline confirmed that [¹¹C]PE2I binds selectively to DAT. In a preliminary study in one human subject the radioactivity ratios of the striatum and substantia nigra to the cerebellum were 10 and 1.8, respectively, at peak equilibrium, which appeared at 40–50 min and 20 min, respectively, after injection. The fraction of the total radioactivity in monkey and human plasma representing unchanged [¹¹C]PE2I was 15–20% at 40 min after injection. The present characterisation of binding in monkey and man suggests that [¹¹C]PE2I is a suitable PET radioligand for quantitative regional examination of DAT in man.