CD45RA+ and CD45RO+ human T cells differ in their functional responses to different forms of mitogenic stimulation. However, little is known about the intracellular signaling events related to the functional differences in these two subsets. In the present study, we examined the ability of different Abs to the TCR/CD3 complex to activate the p21ras proto-oncogene product in CD45RA+ and CD45RO+ T cells. Stimulation with soluble OKT3 anti-CD3 mAb induced activation of p21ras in CD45RA+ cells but not in CD45RO+ cells, although soluble anti-CD3 was able to induce both activation of CD3-associated kinases and IL-2 responsiveness in CD45RO+ cells. Similar results were obtained by using UCHT1, a different anti-CD3 Ab. In contrast, cross-linked anti-CD3 or the combination of phorbol ester and ionomycin were found to activate p21ras equally in both T cell subsets. Our results demonstrate a selective inability of CD45RO+ human T cells to activate p21ras in response to soluble anti-CD3 mAb.