Annexin A2 (AnxA2), a Ca 2+-dependent phospholipid-binding protein, is known to associate with the plasma membrane and the endosomal system. Within the plasma membrane, AnxA2 associates in a Ca 2+ dependent manner with cholesterol-rich lipid raft microdomains. Here, we show that the association of AnxA2 with the lipid rafts is influenced not only by intracellular levels of Ca 2+ but also by N-terminal phosphorylation at tyrosine 23. Binding of AnxA2 to the lipid rafts is followed by the transport along the endocytic pathway to be associated with the intralumenal vesicles of the multivesicular endosomes. AnxA2-containing multivesicular endosomes fuse directly with the plasma membrane resulting in the release of the intralumenal vesicles into the extracellular environment, which facilitates the exogenous transfer of AnxA2 from one cell to another. Treatment with Ca 2+ ionophore triggers the association of AnxA2 with the specialized microdomains in the exosomal membrane that possess raft-like characteristics. Phosphorylation at Tyr-23 is also important for the localization of AnxA2 to the exosomal membranes. These results suggest that AnxA2 is trafficked from the plasma membrane rafts and is selectively incorporated into the lumenal membranes of the endosomes to escape the endosomal degradation pathway. The Ca 2+-dependent exosomal transport constitutes a novel pathway of extracellular transport of AnxA2.