In the previous report, we demonstrated that cerebroside sulfate (CS) and ganglioside GM3 are major acidic glycosphingolipids in rat cochlea. These two glycosphingolipids are also major acidic glycosphingolipids in kidney and furthermore, the both lipids have negative charges. Therefore, the binding activity of one of aminoglycosides, tobramycin to these lipids was studied using [3H] tobramycin. On the thin layer chromatography, tobramycin seemed to bind cerebroside sulfate. On the column chromatograpy using cerebroside sulfate-octyl-Sepharose 4B or GM3 acid-Sepharose, adsorbed tobramycin was eluted with increacing ionic strength in elution buffer. It was assumed that the binding between acidic glycosphingolipids and tobramycin was. of weak ionic bond.