Table 1.

The key attributes of various immobilization methods.

Non-Covalent Immobilization
Glycan type	Advantages	Disadvantages	Examples
Un-modified	Minimal carbohydrate processing	Non-specific binding and orientation; Low MW glycans not stable	High MW polysaccharides on nitrocellulose-coated glass
Linker-modified	Specific binding at linker terminus	Pre-modification required	Biotinylated glycans for avidin-modified surfaces
Covalent Immobilization
Self-assembly
Linker-modified	Facile immobilization process; Compatible with variety of bioanalytical methods	Conjugation with linker required	Glycans fitted with thiol terminated linker for gold surfaces
Unmodified	Activated linker pre-assembled on surface	Potentially non-specific conjugation; May alter carbohydrate structure	Reductive amination or oxime formation to connect immobilized linker
Functional group modifications
Modified with functional group for reaction with surface	Modification at specific carbohydrate position; Controlled presentation; Wide variety of surface materials and formats available	Requires chemical modification of surface and/or sugar	Amine-functionalized sugar conjugated through amide bond formation
Unmodified	No pre-modification of glycan required	Potentially non-specific conjugation; May alter carbohydrate structure	Hydrazine or aminooxy formation at anomeric position; Non-specific linkage to CC
Photochemical
Unmodified	No pre-modification of glycan required	Non-specific binding; Potentially destructive to monosaccharide structure	Surfaces modified with PFPA
Linker-modified	Not destructive; Specific binding	Requires linker installation	Glycan modified with PFPA linker
Cycloaddition
Linker-modified	Amenable to aqueous, bioanalytical conditions; Inkjet compatible	Requires linker installation and compatibly modified surface	CuAAC; Diels-Alder; Staudinger; Strain-promoted
Copolymerization
Natural/Synthetic; Amino-linked; Monomer or allyl-functionalized	Potentially hydrophilic; 3-D cell surface mimic	Requires linker installation; Pre-modification may alter monosaccharide structure	Glycohydrogel; Glyconanopolymers; 3D microarray
Glycosylation
Protected monosaccharides	Synthesis of pure natural and uncommon glycans; Enzymatic synthesis compatible with bioanalytical formats	Time intensive reaction sequences; Chemical synthetic conditions not readily compatible for bioanalysis	Immobilized to activated resin by nucleophilic substitution