Ultrasensitive electrochemical detection of hepatitis C virus core antigen using terminal deoxynucleotidyl transferase amplification coupled with DNA nanowires.

Abstract

Early diagnosis of hepatitis C virus (HCV) infection is essential to prevent disease from spreading and progression. Herein, a novel electrochemical biosensor was developed for ultrasensitive detection of HCV core antigen (HCVcAg) based on terminal deoxynucleotidyl transferase (TdT) amplification and DNA nanowires (DNW). After sandwich-type antibody-antigen recognition, the antibody-conjugated DNA was pulled to theelectrode surface and further extended into a long DNA sequence by robust TdT reaction. Then, large numbers of methylene blue-loaded DNW (MB@DNW) as signal labels are linked to the extended DNA sequence. This results in an amplified electrochemical signal for HCVcAg determination, typically measured at around -0.25V (Ag/AgCl). Under the optimum conditions, the proposed biosensor achieved a wide linear range for HCVcAg from 0.1 to 312.5pg/mL with a low limit of detection of 32fg/mL. Thegood practicality of the biosensor was demonstrated by recovery experiment (recoveries from 98 to 104% with RSD of 2.5-4.4%) and comparison with enzyme-linked immunosorbent assay (ELISA). Given the highlighted performance, the biosensor is expected to act as a reliable sensing tool for HCVcAg determinationin clinics. Schematic representation of the ultrasensitive electrochemical biosensor based on terminal deoxynucleotidyl transferase (TdT) amplification linked with methylene blue-loaded DNA nanowires (MB@DNW), which can be applied to the determination of hepatitis C virus core antigen (HCVcAg) in clinical samples. dTTPs, 2'-deoxythymidine 5'-triphosphate.