Dendritic cells (DC) were cultured from mouse bone marrow (BM) progenitors in low concentrations of granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) (GM^lo DC) by two different protocols. The phenotype and functional properties of these GM^lo DC were compared to those of standard BM‐DC cultures generated in high concentrations of GM‐CSF (GM^hi DC) or in low GM‐CSF plus IL‐4 (GM^lo/IL‐4 DC). An effect of IL‐4 on maturation was observed only at low but not high doses of GM‐CSF. Compared to mature DC, GM^lo DC were phenotypically immature, weak stimulators of allogeneic and peptide‐specific T cell responses, but substantially more potent in presentation of native protein. Immature GM^lo DC were resistant to maturation by lipopolysaccharide, TNF‐α or anti‐CD40 monoclonal antibodies, as the expression of co‐stimulatory molecules was not increased, and stimulatory activity in oxidative mitogenesis was not enhanced. These maturation‐resistant immature GM^lo DC induced T cell unresponsiveness in vitro and in vivo. GM^lo DC also prolonged haplotype‐specific cardiac allograft survival (from 8 days to >100 days median survival time) when they were administered 7 days (but not 3, 14 or 28 days) before transplantation. Our findings may have important implications for future studies in T cell tolerance induction in vivo.