1. Uptake of 3H-25OHD3, 3H-25OHD3-DBP, 125I-holo-DBP and 125I-apo-DBP by LLC-PK1 cells was linearly related to the concentration of each in the culture media. The presence of DBP in the medium significantly reduced the amount of 3H-25OHD3 taken up by cells. 2. Free 25OHD3 and 25OHD3 bound to DBP were both metabolized by the cells to 24,25(OH)2D3 and an unidentified product of apparent lower polarity than 25OHD3. 3. A significant amount of DBP taken up by the LLC-PK1 cells was metabolized to a TCA-soluble form. 4. Uptake of DBP was similar to horseradish peroxidase, but higher than inulin, indicative of a non-specific endocytic mechanism with an adsorptive component. 5. It is suggested that both free circulating 25OHD3 and that derived from lysosomal degradation of 25OHD3-DBP are available for hydroxylation by the kidney.