DNA hybridization biosensor using chitosan-carbon nanotubes composite film as an immobilization platform and [Cu(bpy)(MBZ)_2(H_2O)] (bpy=2,2'-bipyridine, MBZ=p-methylbenzoate) as a novel redox indicator

摘要

In this paper, a new DNA hybridization detection strategy was developed based on the immobilization of capture probe DNA on a chitosan (CS)-carbon nanotubes (CNTs) composite modified glassy carbon electrode (CS-CNTs/GCE) and the use of a copper complex, [Cu(bpy)(MBZ)_2(H_2O)] (bpy=2,2'-bipyridine, MBZ=p-methylbenzoate), as a new redox hybridization indicator. The electrochemical characterization experiments showed that the nanocomposite film of CS-CNTs could effectively immobilize the capture probe DNA and greatly improve the electron-transfer reactions of the electroactive molecules. Electrochemical and fluorescent spectroscopic analysis revealed that the polypyridyl copper complex of [Cu(bpy)(MBZ)_2(H_2O)] bound to DNA via a typical intercalation mode. Surface studies further showed that the copper complex can discriminate between double-stranded and single-stranded DNA that immobilized on the surface of CS-CNTs/GCE. When being utilized as a redox indicator for the detection of hybridization for short DNA species related to phosphinothricin acetyltransferase (PAT), the indicator showed good specificity for recognizing the complementary, three-base mismatched and non-complementary DNA. Under the optimized conditions, the oxidation peak currents of the copper complex enhanced linearly with increases in the concentration of the complementary sequence in the range from 5.0 × 10~(-10) to 1.0 × 10~(-8) M. A detection limit of 5.0 × 10~(-10) M was also obtained based on the constructed DNA biosensor.