Figure 2.

Schematic illustration of the functionalization of the cantilever tip with antibody or WGA. The AFM cantilever tip is first treated with 3‐aminopropyl‐triethoxysilane (APTES) to obtain the amine groups (see Methods) which can react with the NHS group of the PEG linker in the presence of triethylamine. After application of citric acid, the acetal group of the linker converts to aldehyde that subsequently reacts with the amine group of the lysine residue of the anti‐heparan sulfate antibody or WGA. Since both antibody and WGA are coupled to the tip through flexible linkers, they have the possibility to adjust to their ideal orientation for binding to the glycan. In the presence of sodium cyanoborohydride (NaCNBH₃), the proteins are irreversibly linked to the AFM tip. Their density is sufficiently low to allow for single molecule studies [Colour figure can be viewed at wileyonlinelibrary.com]