Flotillins are membrane proteins that form microdomains in the plasma membrane of all mammalian cell types studied to date. They span the evolutionary spectrum, with proteins related to flotillins present in bacteria, fungi, plants and metazoans, which suggests that they perform important, and probably conserved, functions. Flotillins have been implicated in myriad processes that include endocytosis, signal transduction and regulation of the cortical cytoskeleton, yet the molecular mechanisms that underlie flotillin function in these different cases are still poorly understood. In this Commentary, we will provide an introduction to these intriguing proteins, summarise their proposed functions and discuss in greater detail some recent insights into the role of flotillin microdomains in endocytosis that have been provided by several independent studies. Finally, we will focus on the questions that are raised by these new experiments and their implications for future studies.