Phagocytes such as macrophages and dendritic cells (DCs) engulf apoptotic cells to maintain peripheral immune tolerance. However, the mechanism for the recognition of dying cells by phagocytes is not fully understood. Here, we demonstrate that T-cell immunoglobulin mucin-3 (Tim-3) recognizes apoptotic cells through the FG loop in the IgV domain, and is crucial for clearance of apoptotic cells by phagocytes. Whereas Tim-4 is highly expressed on peritoneal resident macrophages, Tim-3 is expressed on peritoneal exudate macrophages, monocytes, and splenic DCs, indicating distinct Tim-mediated phagocytic pathways used by different phagocytes. Furthermore, phagocytosis of apoptotic cells by CD8(+) DCs is inhibited by anti-Tim-3 mAb, resulting in a reduced cross-presentation of dying cell-associated antigens in vitro and in vivo. Administration of anti-Tim-3 as well as anti-Tim-4 mAb induces autoantibody production. These results indicate a crucial role for Tim-3 in phagocytosis of apoptotic cells and cross-presentation, which may be linked to peripheral tolerance.