T lymphocytes with the surface phenotype CD4⁺8⁻ and CD4⁻8⁺ are considered to be representative of functionally mature cells. We show here that adult murine thymus contains a subpopulation of CD4‐8⁺ cells that differ from CD4⁻8⁺ cells found d the periphery in that they do not express the T cell receptor‐associated CD3 molecular complex. Such CD3⁻4⁻8⁺ thymocytes are cortisone sensitive and rapidly cycling in situ. Furthermore, in contrast to mature T cells, most CD3⁻4⁻8⁺ thymocytes express low levels of CD5 and high levels of the B2A2 antigen. CD3⁻4⁻8⁺ thymocytes fail to respond to a variety of mitogenic stimuli in vitro but do give rise upon short‐term culture to CD4⁺8⁺ cells. It is suggested that CD3⁻4⁻8⁺ thymocytes represent a transitional stage of thymus differentiation between the CD4⁻8⁻ and CD4⁺8⁺ compartments.