Recent studies have focused on the ability of tocopherols to regulate gene expression. For such experiments, the methodology used to deliver molecules to the cells is crucial and could lead to different results depending on the vehicle used. The objective of the present study was to compare commonly used tocopherol vehicles (ethanol, BSA and mixed micelles) in terms of toxicity, stabilization of tocopherols, uptake efficiency of tocopherols by cells and effect on gene expression. Lactate dehydrogenase measurements did not reveal cytotoxicity of any of the tested vehicles. Tocopherol recovery measurements showed that approximately 80% of the tocopherol was lost in ethanolic solutions, while only approximately 30% and 10% were lost in BSA and mixed micelles, respectively. After 24 h incubation, Caco-2 cell monolayers treated with mixed micelles exhibited the highest alpha-tocopherol intracellular concentrations (5-times those measured with the two other vehicles). Similar results were obtained with gamma-tocopherol. Vehicles, except mixed micelles that activate the FXR/bile acids signalling pathway, did not affect expression of nuclear receptors involved in lipid metabolism or their target genes. This study establishes mixed micelles as the best vehicle to deliver tocopherols to intestinal cell monolayers in culture.