Investigation of the effects of 1,25(OH)2D3 and 24,25(OH)2D3 on the proliferation and differentiation of the human myelomonocytic cell line U937 has been complemented with studies of the effect of the same metabolites on the number of nuclear receptors for 1,25(OH)2D3. Both 1,25(OH)2D3 and 24,25(OH)2D3 inhibit the proliferation of U937 cells in a dose-dependent manner. The concentrations of 24,25(OH)2D3 required to produce this effect were 100-times greater than those of 1,25(OH)2D3. Inhibition of proliferation was associated with increased expression of the CD14 and 200 kDa 63D3 antigens thus confirming differentiation of U937 towards a more mature cell type. Studies of the nuclear receptor for 1,25(OH)2D3 showed that pre-treatment of the cells with 1,25(OH)2D3 resulted in an apparent 40% decrease in the number of detectable 1,25(OH)2D3 receptors as compared to control U937 cells. This is due to the fact that the 1,25(OH)2D3 binds to U937 cell nuclei during culture and thus blocks the subsequent binding of radiolabelled 1,25(OH)2D3 used to measure the number of 1,25(OH)2D3 receptors. Measurement of the binding of unlabelled 1,25(OH)2D3 by radioimmunoassay indicated that pre-treatment of the cells with 1,25(OH)2D3 increased the capacity of U937 to bind the hormone, although measurement of these receptors by whole cell assay was prevented by the binding of 1,25(OH)2D3 itself. This effect was not observed with 24,25(OH)2D3 which was more easily displaced from binding sites by radiolabelled 1,25(OH)2D3 and it appears to act through low affinity binding to the 1,25(OH)2D3 receptor.