Oxime chemistry-mediated covalent capturing on electrode surface with guanidinium recognition and application for aldolase activity assay

A new strategy to fabricate electrochemical biosensor is proposed based on the covalent capturing of molecules on electrode interface through oxime chemistry and the loading of signal probes via guanidinium-phosphate electronic interaction with a “covalent-like” stability. Taking aldolase as an example, enzyme analysis can be achieved through the covalent loading of the catalyzed products on the aminooxy modified gold electrode surface and subsequent immobilization of guanidinium functionalized sliver nanoparticles. Different from adolase substrate, the hydrolyzed products contain both carbonyl groups and phosphate acid groups in their structures. The carbonyl groups can chemoselectively react with aminoxy groups on the electrode surface to form oxime, resulting in the exposure of phosphate groups. Subsequently, signal probe can be loaded onto electrode surface through guanidinium recognition, giving a electrochemical current value. So the number of enzyme catalyzed products determines the amount of signal probes on the electrode surface, and aldolase activity can be analyzed. For the proposed method, the chemoselective oxime ligation and highly stable guanidinium recognition improve the accuracy, selectivity and stability of enzyme analysis. The established method may be extended to the analysis of other related substances containing carbonyl groups.