Generation of effective CTL responses is the goal of many vaccination protocols. However, to what extant T cell precursor frequencies will generate a CD8(+) CTL response has not been elucidated properly. In this study, we employed a model system, in which naive CD4(+) and CD8(+) T cells derived from ovalbumin (OVA)-specific TCR transgenic OT II and OT I mice were used for adoptive transfer into wild-type, Ia(b-/-) gene knockout and transgenic RIP-mOVA mice, and assessed OVA-pulsed DC (DCOVA)-stimulated CD8(+) CTL responses in these mice. We demonstrated that (i) a critical threshold exists above which T cells precursor frequency cannot enhance the CTL responses in wild-type C57BL/6 mice, (ii) increasing CD8(+) T cell precursors is required to generate CTL responses but with functional memory defect in absence of CD4(+) T cell help, and (iii) increasing CD4(+) and CD8(+) T cell precursors overcomes immune suppression to DCOVA-stimulated CD8(+) CTL responses in transgenic RIP-mOVA mice with OVA-specific self immune tolerance. Taken together, these findings may have important implications for optimizing immunotherapy against cancer. (c) 2007 Elsevier Inc. All rights reserved.