Carbon Nanotubes Labeled with Aptamer and Horseradish Peroxidase as a Probe for Highly Sensitive Protein Biosensing by Postelectropolymerization of Insoluble Precipitates on Electrodes

Carbon nanotubes (CNTs) labeled with aptamer and horseradish peroxidase (HRP) were used as a probe to amplify the impedimetric sensing of the aptamer–protein (with thrombin as the model) interaction. The HRP-biocatalyzed oxidation of 3,3-diaminobenzidine (DAB) in the presence of H₂O₂ and the postelectropolymerization of insoluble precipitates produced on the electrode supports were used as a signal amplification route for the sensing process. Thrombin was sensed by aptamer 1 immobilized on a glassy carbon electrode. The multiwalled CNT–aptamer 2–HRP probe was linked to the aptamer 1–thrombin complex through the thrombin–aptamer 2 interaction. The postelectropolymerization of biocatalyzed precipitates of DAB on the electrode greatly increased the electron-transfer resistance at the electrode–solution interface. Cyclic voltammetry and electrochemical impedance spectroscopy were employed to follow the stepwise fabrication of the aptasensor and impedimetric detection of thrombin. Thrombin concentration as low as 0.05 pM could be detected by this method. In addition, the proposed impedimetric aptasensor exhibits good sensitivity (5195 Ω decade^–1), selectivity, and reproducibility. The aptasensor also has acceptable recovery for thrombin detection in complex protein sample