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We designed and synthesized three tetravalent sialo-glycoclusters that had different separations between the terminal sialic acids and the linking carboxy groups of the ethylene glycol bis(β-aminoethyl ether)-N,N,N′,N′-tetraacetate scaffold to serve as ligands for the sialic acid-binding lectin Sambucus sieboldiana agglutinin (SSA). The interaction between each glycocluster and SSA was characterized by hemagglutination inhibition, quantitative precipitation, and double-diffusion assays. For the precipitation assays, the precipitin curves indicated that the ligands and SSA bound in either a 1:1 or a 1:2 ratio, i.e., stoichiometrically. The strong interactions of these sialo-glycoclusters with SSA could be ascribed to a combination of multivalency and spacer effects. We also assessed the nature of the ligand–SSA complexes by isothermal titration calorimetry and dynamic light scattering. The results of those experiments …