CD83 is the major surface marker identifying mature dendritic cells (DCs). In this study, we report the generation of reporter mice expressing EGFP under the control of the CD83 promoter. We have used these mice to characterize CD83 expression by various immune system cell types both in vivo and ex vivo and under steady-state conditions and in response to stimulation with a Toll-like receptor (TLR) ligand. With those mice we could prove in vivo that the CD83 promoter is highly active in all DCs and B cells in lymphoid organs. Interestingly, this promoter activity in B cells mainly depended on the stage of development, is up-regulated in the late pre-B cell stage, and was maintained on a high level in all peripheral B cells. We also confirmed that CD83 in those cells is mainly intracellular but is up-regulated after TLR stimulation. Otherwise, CD83 promoter activity in T cells seemed to depend on stimulation and could be found mainly in CD4⁺CD25⁺ and CD8⁺CD25⁺ T cells and in CD4⁺ and CD8⁺ memory cells. In addition, we identified the murine homologues of the human CD83 splice variants. In contrast to those in human, those extremely rare short transcripts were never found without the expression of the highly dominant full-length form. So, the murine CD83 surface expression is mainly regulated posttranslationally in vivo. Our CD83 reporter mice represent a useful mouse model for monitoring the activation status and migration of DCs and lymphocytes under various conditions, and our results provide much needed clarification of the true nature of CD83 promoter activity.