It is well established that naive cells can be converted by TGF-β into CD4(+)CD25(+) regulatory T (Treg) cells with therapeutic potentials. Likewise, it is shown that all-trans retinoic acid (ATRA) can greatly enhance TGF-β-induced Treg conversion, a phenomenon which has mainly been studied in C57BL/6 mice. Here we show that, although purified naive cells are highly susceptible to Treg generation, total CD4(+) T-cell populations from different mouse strains display significantly different sensitivities to TGF-β/ATRA-induced Treg conversion. The resistance of "non-responder" strains is associated with an enhanced production of IL-4 by memory T cells as well as an increased sensitivity of naive T cells to the action of IL-4. Importantly, neutralization of IL-4 overcomes the differences, thereby enabling TGF-β/ATRA to generate large numbers of functional Treg cells from total CD4(+) T cells in a consistent manner across different mouse strains. Likewise, blockade of IL-4 significantly enhances TGF-β/ATRA-induced Treg conversion from human naive T cells in the presence of memory cells. These results show that the inherent resistance of "non-responder" mouse strains to Treg conversion induced by TGF-β and ATRA can be reverted by neutralization of IL-4 and that inhibiting the action of IL-4 is beneficial or even necessary for consistent inducible Treg generation.