Memory CD8+ T cells from mice previously primed with alloantigen (alloAg) can respond in vitro to IL-2 and purified class I alloAg presented on microspheres, while no response can be detected using cells from naive mice. Similar results have been obtained using cells from OT-1 mice expressing a transgenic TCR that is specific for OVA 257–264 (SIINFEKL) peptide bound to H-2K b. A population of resting memory cells (defined on the basis of low forward scatter and CD44 high, Ly-6C+, CD25−, CD69− surface phenotype) that is present in the OT-1 mice exhibits a substantially higher sensitivity to Ag-stimulation than do naive cells (CD44 low, Ly-6C−) expressing the same TCR. CD44 high cells respond vigorously to H-2K b immobilized on microspheres and pulsed with peptide, while CD44 low cells respond weakly and only at high class I density and peptide concentration. The Ag-presenting surface only has ligands for TCR and CD8 (class I and peptide), thus ruling out the possibility that differences are due to ligand binding by other adhesion or costimulatory receptors that are expressed at high levels on the memory cells. Experiments using anti-TCR mAb as the stimulus and coimmobilized non-Ag class I as a ligand for CD8 suggest that the difference between naive and memory cells may be at the level of stimulation through the TCR. Thus, in addition to expressing increased levels of adhesion receptors that may enhance responses to Ag on APCs, memory CD8+ T cells appear to be intrinsically more sensitive than naive cells to stimulation through the TCR/CD8 complex.