The production of dendritic cells (DC) from haemopoietic progenitors maintained in long term stroma-dependent cultures (LTC) of spleen or bone marrow (BM) occurs independently of added granulocyte/macrophage colony stimulating factor (GM-CSF). The possibility that cultures depend on endogenous GM-CSF produced in low levels was tested by attempting to generate LTC from spleen and BM of GM-CSF-/- mice. Multiple cultures from GM-CSF-/- and wild type mice were established and compared for cell production. GM-CSF-/- LTC developed more slowly, but by 16 weeks produced cells resembling DC in numbers comparable to wild type cultures. LTC maintained distinct populations of small and large cells, the latter resembling DC. Cells collected from GM-CSF-/- LTC were capable antigen presenting cells (APC) for T cell stimulation and morphologically resembled DC. Large cells expressed the CD11b, CD11c, CD86, 33D1 and Dec-205 markers of DC. Addition of GM-CSF to GM-CSF-/- LTC increased the proportion of large, mature DC present in culture. Stromal cells from GM-CSF-/- LTC could support the differentiation of DC from early progenitors maintained in LTC without addition of GM-CSF. However, GM-CSF is not a critical factor in the in vitro generation of DC from progenitors. It can, however, substitute for stromal cells in increasing the survival of mature DC.