The acute inflammatory response requires that circulating leukocytes adhere to, and then migrate through, the vascular wall at the site of injury or infection. Several receptors have been implicated in this adhesion and migration process, including the selectins, a family of carbohydrate-binding proteins. The ligand for one of these proteins, E-selectin (LECAM-2, ELAM-1) has been described by several groups to contain a polylactosamine structure bearing a terminal sialic acid residue and at least one fucose residue. We report here a more detailed investigation into the minimum structural requirements for carbohydrate recognition by E-selectin. Using both direct binding and inhibition studies we demonstrate that the sialyl Lewisx tetrasaccharides Sia(alpha 2-3)Gal(beta 1-4)[Fuc(alpha 1-3)]GlcNAc, and Sia(alpha 2-3)Gal(beta 1-4)[Fuc(alpha 1-3)]Glc are the smallest oligosaccharides recognized by the lectin. In addition, an oligosaccharide containing the sialyl Lewisa epitope is also recognized, but less avidly. We propose a structural model of functional groups necessary for recognition by E-selectin, based on these data and additional experiments on modifications of sialic acid and the reducing terminal saccharide.