Detecting Circulating Anti-Glycan Antibodies with a Printed Glycan Array

Begin with a functionalized glycochip featuring printed glycan sub-arrays organized into blocks of arrays.

Each spot in the sub-array represents a distinct glycan — a carbohydrate-based polymer with variations in monosaccharide composition, chain length, and branching pattern.

Treat the glycochip with a blocking buffer to block non-specific interaction sites.

Transfer the glycochip and introduce diluted mouse serum in a surfactant-supplemented buffer to minimize antibody aggregation. Incubate under agitation. 

Agitation promotes uniform serum distribution, facilitating selective binding of anti-glycan antibodies to specific glycans based on distinct molecular characteristics.

Wash with decreasing concentrations of a surfactant-containing buffer to disturb non-specific binding and rinse.

Overlay the glycochip with anti-mouse biotin-conjugated secondary antibodies, interacting with pre-bound anti-glycan antibodies and wash.

Introduce fluorophore-labeled streptavidin, binding to biotinylated secondary antibodies. Wash and remove excess solution.

Scan the glycochip and observe for distinct fluorescence spots at various positions suggesting glycan-specific circulating antibodies in serum.