Langerhans cells (LCs) are the main population of antigen-presenting cells lining the epidermis and stratified mucosal epithelia (1). Therefore, they play an important role in the first line of defense against invading pathogens. Upon capture of these pathogens, LCs subsequently migrate to the lymph nodes where they present pathogen-derived antigens to T cells to initiate an adaptive immune response. During this migration, LCs up-regulate cell surface marker HLA-DR and co-stimulatory molecules, while the LC-specific C-type lectin Langerin is down-regulated (reviewed in Refs. (2,3)). In the epidermis, LCs are the only cell population expressing CD1a and this marker is therefore extremely useful to isolate LCs from epidermis (4). Here we discuss a method to isolate human primary LCs from the epidermis in an as immature state as possible. The use of immature LCs is especially important in the investigation of the function of these cells, since few acceptable LC models are available. Immature LCs can be used to further elucidate the function of LCs in pathogen interactions and adaptive immunity.