Electrochemical immunosensor based on an azo compound for thyroid-stimulating hormone detection

An electrochemical immunosensor based on the electroactive azo compound (E)-5-[(4-dodecyloxyphenyl)diazenyl]isophtalic acid was developed for thyroid-stimulating hormone (TSH) detection. This organic molecule was employed as a multifunctional platform actuating simultaneously as redox probe and active site for covalent immobilization of the monoclonal anti-TSH antibody (ab-TSH) via 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide/N-hydroxysuccinimide (EDC/NHS) chemistry by means of the carboxylic groups present in the azo compound. The functioning principle of the immunosensor is the decrease in electrochemical response arising from the azo compound film deposited on the surface of a glassy carbon electrode (GCE) due to the presence of TSH antigen, which interacts specifically with ab-TSH antibodies immobilized on the electrode surface. The signal inhibition is proportional to the amount of antigens present in the analyzed sample. The electrochemical performance was studied by cyclic and square-wave voltammetry and electrochemical impedance spectroscopy. The immunosensor surface was characterized by electron microscopy. Under optimal conditions, a linear range of 0.2 to 20.0 μIU mL− 1 with a calculated detection limit of 0.04 μIU mL− 1 was achieved. The evaluation of some potential interfering compounds (glucose, ascorbic acid, uric acid and creatine) resulted in a low degree of interference (< 10%) and the immunosensor presented a good inter-assay precision (< 5%). The proposed immunosensor was successfully applied for TSH determination in simulated blood serum showing good accuracy and sensitivity.