QCM-D and SE were used to study the binding between STxB or CTB and the different glycocalyx models built using HA-B, (HA-g-Lac L )-B, (HA-g-Gb3)-H6, (HA-g-Gb3)-B or (HA-g-Le x )-B. These lectins showed selective binding to the brushes containing their target glycans: Le x trisaccharide was recognized only by CTB, while films with Gb3 bound only to STxB. The StxB-Gb3 interaction (Kd = 1.8 ± 0.2 µM) was enhanced 1000-fold relative to the highest affinity monovalent Gb3-oligosaccharide interaction, yet this is still 500-fold lower affinity than reported for STxB binding to the Gb3 glycosphingolipid in a membrane. This is perhaps not surprising when one considers that the STxB protein architecture has evolved to have all its binding sites on one flat face of the protein which is optimal for binding to a surface rather than to a 3D structure like our glycocalyx model. Nonetheless, at saturation the Gb3:STxB pentamer ratio was >10 (Figure ), which could be consistent with all of the StxB 'site 1' and 'site 2' binding sites being occupied. However, other multivalent Gb3 ligands only engage the higher affinity 'site 2', and so the data could also result from only partial saturation of the Gb3 ligand groups.