Functionalization of cantilever tips with nucleotides by the phosphoramidite method

In atomic force microscopy (AFM) a sharp cantilever tip is used to scan surfaces at the atomic level. One further application is force spectroscopy, in which force-distance curves between binding partners located on the cantilever and substrate surface are determined. This requires specifically immobilized molecules. Herein we describe the covalent binding of single adenosine and thymidine nucleotides on an amino-PEGylated cantilever tip by the phosphoramidite method. Force-distance curves between these cantilever tips and gold surfaces were recorded. The rupture forces of the coordination bond between the primary amine of adenosine and the undercoordinated gold atoms were determined to be 145 pN, which is in agreement with previously published data. The force-distance curves of thymidine-functionalized tips did not show rupture events, because this nucleotide does not possess a primary amine function. Nucleotide-functionalized tips could aid in the understanding of binding mechanisms of nucleotide binding molecules such as polymerases immobilized on surfaces or membranes.