Human Langerhans cells (LC) can be generated ex vivo from hematopoietic precursor cells in response to cytokines and cell-membrane associated ligands. These in vitro differentiation models provided mechanistic insights into the molecular and cellular pathways underlying the development of this unique, epithelia-associated dendritic cell subset. Notably, the human epidermal microenvironment is fully sufficient to induce LC differentiation from hematopoietic progenitors. Hence, dissecting the molecular characteristics of the human epithelial/epidermal LC niche, and testing defined ligands for their capacity to induce LC differentiation, led to a refined molecular model of LC lineage commitment. During epidermal ontogeny, spatially and temporally regulated availability of TGF-β family members cooperate with other keratinocyte-derived signals, such as E-cadherin and Notch ligands, for instructing LC differentiation. In this review, we discuss the signals known to instruct human hematopoietic progenitor cells and myelomonocytic cells to undergo LC lineage commitment. Additionally, the current methods for generation of large numbers of human LC-like cells ex vivo in defined serum-free media are discussed.