Prion diseases ale fatal neurodegenerative disorders resulting from structural conversion of the cellular isoform of PrPC to the infectious scrapie isoform PrPSc. It is believed that such structural alteration may occur within the internalization pathway However, there is no direct evidence to support this hypothesis Employing quantum dots (QDs) as a probe, we have recorded a real-time movie demonstrating the process of prion internalization in a living cell for the first time The entire internalization process can be divided into four discrete but connected stages In addition, using methyl-beta-cyclodextrin to disrupt cell membrane cholesterol, we show that lipid rafts play an important role in locating cellular PrPC to the cell membrane and in initiating PrPC endocytosis (C) 2010 Elsevier Inc All rights reserved