Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a member of the TNF family of cytokines. TRAIL has gained much attention because of its ability to preferentially kill tumor cells with no apparent toxic side effects. Recently, different TRAIL receptor agonists, including TRAIL itself and various agonistic monoclonal antibodies against the two apoptosis-inducing human TRAIL receptors, have been developed as novel cancer therapeutics and are currently under investigation in clinical trials. However, the mechanisms by which TRAIL mediates its selective antineoplastic activity are still not well understood. In addition to playing a role in cancer immune surveillance and tumor suppression, TRAIL has been associated with immune homeostasis, inflammatory diseases, and autoimmunity. In light of the multifunctional role of TRAIL in mediating various pathologic conditions and the potential benefits of TRAIL-based therapies, the study of the physiologic significance of TRAIL is of great importance. Here, we describe a syngeneic system for the characterization of the in vivo function of TRAIL. By use of this model, in which the full-length murine TRAIL protein is overexpressed in the hematopoietic cells of wild-type mice, the in vivo tumoricidal activity of TRAIL overexpression can be studied on syngeneic murine tumor cell challenge, and the potential toxicity of TRAIL protein to normal tissues can also be analyzed.