L-Selectin, a leukocyte adhesion molecule, mediates leukocyte rolling on the endothelium and plays a critical role in leukocyte recruitment at inflammatory sites as well as in lymphocyte homing. We have previously shown that L-selectin reactive chondroitin sulfate and heparan sulfate proteoglycans (HSPGs) are both expressed in the distal tubules of the kidney and that versican is one of the chondroitin sulfate-type ligands. In the present study, we characterized the heparan sulfate-type ligand(s) in more detail. The molecular sizes of HSPGs were approximately 600 kDa with core protein sizes of 160 and 180 kDa. Western blotting analysis showed that L-selectin reactive HSPGs were neither agrin nor perlecan, major basement membrane HSPGs in the kidney. The binding to L-selectin was mediated by the lectin domain of L-selectin in a Ca2+-dependent manner and required heparan sulfate side chains, but not sialic acid. To our knowledge, this is the first biochemical characterization of the L-selectin reactive heparan sulfate proteoglycan(s) in the distal tubules of the kidney.