The sulfate transport in AH-66 hepatoma ascites cells was examined under various controlled conditions using 35SO42- as a tracer. The sulfate efflux rate was dependent on temperature, pH and anion species of the cell suspending medium. The efflux rate became saturated as the concentration of extracellular anions was increased. The efflux of anion was inhibited by some chemical reagents specifically reactive with amino or sulfhydryl groups. The results obtained in this study suggest that sulfate anions were transported by a facilitated transport system(s), and that some membrane protein(s) is involved in the anion transport system(s) of AH-66 cells. Both amino and sulfhydryl groups are thought to play a determinant role at the sulfate transport site in AH-66 cells.