New "clickable" polymeric coating for glycan microarrays

The interaction of carbohydrates with a variety of biological targets, including antibodies, proteins, viruses and cells are of utmost importance in many aspects of biology. Glycan microarrays are increasingly used to determine the binding specificity of glycan-binding proteins. In this study, a novel slide is reported for the fabrication of glycan arrays that combines the higher sensitivity of a layered Si-SiO<inf>2</inf> with a novel approach to form a polymeric coating easily modifiable by subsequent click reaction. The alkyne-containing copolymer, adsorbed from an aqueous solution, produces a coating by a single step procedure and serves as a soft, tridimensional support for the oriented immobilization of carbohydrates via azide/alkyne Cu(I) catalyzed "click" reaction. The equilibrium and kinetics parameters of the interaction of Concanavalin A with eight synthetic glycans were determined using fluorescence microarray and Reflective Phantom Interface (RPI), a recently proposed optical label-free detection approach. The enhancement of fluorescence provided by the Si-SiO<inf>2</inf> slides enabled to extend the limit of detection at lower surface densities of lectins, in turn enabling the study of the interaction for a wide range of glycans surface density. Equilibrium dissociation constants of a few nM were extracted for multivalent glycan-lectin binding, mimicking the conditions of biological membranes, whereas hundreds of nM were observed at the lower glycan surface densities.