The study identifies Tim4 as a phosphatidylserine (PS) receptor involved in the engulfment of apoptotic cells by macrophages. Tim4, a type I transmembrane protein, was identified through the use of a library of hamster monoclonal antibodies against mouse peritoneal macrophages. The antibody Kat5-18, which strongly inhibited PS-dependent engulfment, was found to recognize Tim4. Tim4 was expressed in various mouse tissues, including spleen, lymph nodes, and fetal liver, and bound apoptotic cells by recognizing PS via its immunoglobulin domain. Expression of Tim4 in fibroblasts enhanced their ability to engulf apoptotic cells. In vivo, administration of anti-Tim4 monoclonal antibody to mice blocked the engulfment of apoptotic cells by thymic macrophages and induced the development of autoantibodies. Among other Tim family members, only Tim1 specifically bound PS. Tim1- and Tim4-expressing B cells bound exosomes via PS, and exosomes stimulated the interaction between Tim1 and Tim4. These findings suggest that Tim4 and Tim1 are key phosphatidylserine receptors for the engulfment of apoptotic cells and may also play a role in intercellular signaling involving exosomes.The study identifies Tim4 as a phosphatidylserine (PS) receptor involved in the engulfment of apoptotic cells by macrophages. Tim4, a type I transmembrane protein, was identified through the use of a library of hamster monoclonal antibodies against mouse peritoneal macrophages. The antibody Kat5-18, which strongly inhibited PS-dependent engulfment, was found to recognize Tim4. Tim4 was expressed in various mouse tissues, including spleen, lymph nodes, and fetal liver, and bound apoptotic cells by recognizing PS via its immunoglobulin domain. Expression of Tim4 in fibroblasts enhanced their ability to engulf apoptotic cells. In vivo, administration of anti-Tim4 monoclonal antibody to mice blocked the engulfment of apoptotic cells by thymic macrophages and induced the development of autoantibodies. Among other Tim family members, only Tim1 specifically bound PS. Tim1- and Tim4-expressing B cells bound exosomes via PS, and exosomes stimulated the interaction between Tim1 and Tim4. These findings suggest that Tim4 and Tim1 are key phosphatidylserine receptors for the engulfment of apoptotic cells and may also play a role in intercellular signaling involving exosomes.