Animal tests, such as the local lymph node assay (LLNA), are the gold standard for assaying skin-sensitizing potential. However, because of concerns about animal welfare, extensive research has been conducted on the use of various cell lines, such as human leukemia cells, for in vitro assays of skin-sensitizing potential, but such assays have not replaced animal tests as stand-alone assays. Because Langerhans cells—a type of dendritic cell—are the main antigen-presenting cells in the epidermis and because they play a central role in the induction of allergic skin disorders, these cells may be useful for skin-sensitizing-potential assays. Here, we investigated the utility of the murine dendritic cell line DC2.4 for in vitro assay of the skin-sensitization potential of 2,4-dinitrochlorobenzene (DNCB), 2-mercaptobenzothiazole (MBT), and α-hexyl cinnamaldehyde (HCA), which are categorized as extremely, moderately, and weakly sensitizing, respectively, on the basis of LLNA results. DC2.4 cell viability decreased dose-dependently with increasing concentration upon treatment with each of the compounds for 24 hr; the DNCB, MBT, and HCA concentrations that resulted in 75% cell viability were 6.07, 120.14, and 118.70 μg/mL, respectively. At nontoxic concentrations (concentrations less than the 75% cell viability concentrations), these compounds dose-dependently upregulated the expression of both CD86 and CD54 on the surface of DC2.4 cells. Their potency decreased in the order DNCB > MBT > HCA, which agrees with the order indicated by the LLNA. These results suggest that DC2.4 cells may be a viable replacement for human leukemia cells in in vitro assays of skin-sensitization potential.