Interactions between T cells and antigen-presenting cells are the first step in the induction of an adaptive immune response. Here, we show that CD6 and its ligand ALCAM are actively recruited to the antigen-induced DC-T cell contact zone. Moreover, ALCAM-blocking antibodies interfere with DC-T cell conjugate formation, demonstrating that CD6-ALCAM binding is essential for stable T cell-APC contact. We now demonstrate that besides their role in establishing initial contacts, CD6-ALCAM interactions are also required during the proliferative phase of the T cell response; the presence of CD6-blocking antibodies or recombinant ALCAM-Fc proteins results in a strong and sustained inhibition of T cell proliferation. Furthermore, simultaneous crosslinking of CD6 and CD3 induces enhanced proliferation and transcriptional activity to a similar level as observed after CD3 and CD28 co-crosslinking, demonstrating that CD6 is an important co-stimulatory molecule. The stability of ALCAM-CD6 binding, which contrasts with transient homotypic ALCAM-ALCAM interactions, further supports the long-lasting effects observed on T cell proliferation. Taken together, we demonstrate that CD6 and ALCAM form a key adhesive receptor-ligand pair that is not only involved in early DC-T cell binding but also in sustaining DC-induced T cell proliferation long after the initial contact has been established.