Triplex DNA-based Bioanalytical Platform for Highly Sensitive Homogeneous Electrochemical Detection of Melamine

Melamine detection has attracted much attention since the discovery of the damage of melamine to human health. Herein, we have developed a sensitive homogeneous electroanalytical platform for melamine detection, which is relied on the formation of triplex molecular beacon integrated with exonuclease III (Exo III)-mediated signal amplification. The formation of triplex molecular beacon was triggered by the recognition and incorporation of melamine to the abasic (AP) site contained in the triplex stem. The stem of the triplex molecular beacon was designed to have a protruding double-strand DNA, which can be recognized and hydrolyzed by Exo III for releasing methylene blue (MB)-labeled mononucleotide. These released MB molecules exhibit high diffusivity toward indium tin oxide electrode with negative charge, thus producing a significantly increased electrochemical response. Taking advantages of the high binding affinity of the DNA triplex structure containing AP sites towards melamine and the unique features of Exo III, this sensing platform is capable for sensitive and selective melamine assay with a detection limit as low as 8.7 nM. Furthermore, this strategy shows good applicability for melamine assay in real samples. Therefore, this strategy broadens the application of triplex DNA and presents a new method for sensitive detection of melamine.