Anti‐CD4 monoclonal antibodies are potential therapeutic agents for the prevention of autoimmune disease and treatment of rejection after organ transplantation and are capable of both restoring tolerance to self‐antigens and inducing tolerance to antigens introduced under the cover of the antibody therapy in vivo. Tolerance to donor alloantigens can be induced in vivo by administering donor alloantigen in combination with either depleting (YTA 3.1) or nondepleting (YTS 177) anti‐CD4, 28 days before heart transplantation in the mouse. The effect of anti‐CD4 on proximal T‐cell receptor (TCR) signalling pathways and proliferation was investigated in vitro and in vivo in the presence and absence of YTA 3.1 or YTS 177. Anti‐CD4 was found to perturb proximal signalling events upon TCR/CD3 ligation, resulting in reduced tyrosine phosphorylation of Zap‐70 and LAT (linker for activation of T cells) and reduced association of tyrosine‐phosphorylated LAT with lck. This ultimately resulted in severely reduced proliferation of the responding CD4⁺ T cells. The signalling profile of the anti‐CD4‐treated cells resembled that of anergic T cells. This could be a result of a common mechanism involving perturbation in the formation of the central supramolecular activation cluster of the immunological synapse by impaired recruitment of CD4 and CD28, thereby resulting in severely reduced lck activation.