The phenotypic and functional characteristics of Ag-specific memory CD4+ lymphocytes are poorly defined. To examine the properties and cytokine responsiveness of these cells, we have developed an adoptive transfer system using in vitro-activated T cells expressing the DO.11 transgenic TCR specific for OVA(323-339)+ I-Ad. In vitro-activated DO.11 CD4+ cells exhibit comparable survival patterns at 1, 6, and 10 wk after adoptive transfer, indicating that a stable population of memory cells has been generated. In the absence of Ag, previously activated T cells survive longer than their naive counterparts in vivo, rapidly revert to a partially naive phenotype, and maintain their effector cytokine profile. The DO.11 CD4+ memory cells are capable of proliferating in response to IL-2 and IL-4, while naive DO.11 CD4+ cells exhibit no such proliferative responses.