Structurally diverse glycans are expressed by all animate beings and exert diverse biological functions through specific interactions with glycan binding proteins (GBPs). In humans, glycan-GBP interactions are implicated in many disease-relevant processes in development, infection and immune response to bacterial and viral pathogens. Recent progress in chemical synthesis, including automated glycan assembly, has facilitated access to complex glycans that cannot be isolated from biological material. Glycan immobilization on microarrays allows rapid, multiplexed glycan-GBP interaction studies to reveal biological functions. Synthetic glycan microarrays have enabled, for instance, the identification of glycan ligands for lectins, the definition of vaccine antigens, revealed viral glycan receptors and can serve as diagnostic tools for human disease. Here, we describe the methods to fabricate custom glycan microarrays that are used to examine glycan-GBP binding specificities. Conjugation-ready synthetic glycans are covalently attached to microarray surfaces through nucleophilic linker moieties. Microarrays are incubated with GBPs, and binding events are quantitatively detected by fluorescent signals. These methods are readily adaptable to a multitude of purposes from basic research to biomedical applications.
Keywords: Automated glycan assembly (AGA); Glycan; Glycan binding protein (GBP); Glycan–GBP interaction; Microarray; Oligosaccharide.