Highly sensitive electrochemical impedance spectroscopic detection of DNA hybridization based on Aunano–CNT/PANnano films

A polyaniline nanofibers (PANnano)/carbon paste electrode (CPE) was prepared via dopping PANnano in the carbon paste. The nanogold (Aunano) and carbon nanotubes (CNT) composite nanoparticles were bound on the surface of the PANnano/CPE. The immobilization and hybridization of the DNA probe on the Aunano–CNT/PANnano films were investigated with differential pulse voltammetry (DPV) and cyclic voltammetry (CV) using methylene blue (MB) as indicator, and electrochemical impedance spectroscopy (EIS) using [Fe(CN)6]3−/4− as redox probe. The voltammetric peak currents of MB increased dramatically owing to the immobilization of the probe DNA on the Aunano–CNT/PANnano films, and then decreased obviously owing to the hybridization of the DNA probe with the complementary single-stranded DNA (cDNA). The electron transfer resistance (Ret) of the electrode surface increased after the immobilization of the probe DNA on the Aunano–CNT/PANnano films and rose further after the hybridization of the probe DNA. The remarkable difference between the Ret value at the DNA-immobilized electrode and that at the hybridized electrode could be used for the label-free EIS detection of the target DNA. The loading of the DNA probe on Aunano–CNT/PANnano films was greatly enhanced and the sensitivity for the target DNA detection was markedly improved. The sequence-specific DNA of phosphinothricin acetyltransferase (PAT) gene and the polymerase chain reaction (PCR) amplification of nopaline synthase (NOS) gene from transgenically modified beans were determined with this label-free EIS DNA detection method. The dynamic range for detecting the PAT gene sequence was from 1.0 × 10−12 mol/L to 1.0 × 10−6 mol/L with a detection limit of 5.6 × 10−13 mol/L.