An Electrochemical Biosensor for the Sensitive Detection of Hepatitis C Virus in Unpurified Polymerase Chain Reaction Amplified Real Samples based on Peptide Nucleic Acid and Double-stranded DNA Hybridization

Abstract

Several studies show that acute infections with hepatitis C virus (HCV) frequently progresses to chronic diseases, eventually can lead to liver cirrhosis and hepatocellular carcinoma. Thus, development of simple and reliable HCV detection methods is in demand. The present paper describes electrochemical detection of polymerase chain reaction (PCR)-amplified Core/E1 encoding cDNA corresponding to hepatitis C virus (573 bp size) directly in double stranded form without any purification, pre-treatment and the need for denaturation of the target. The biosensor relies on the hybridization between self-assembled cysteine conjugated 20-mer peptide nucleic acid (PNA) oligomer probe and complementary ds-PCR products to form PNA-ds-PCR hybrid. The extent of hybridization between the probe and target sequences was determined by using differential pulse voltammetric signal of methylene blue (MB) as the hybridization indicator. In order to improve biosensing performance, the effect of various factors was investigated. The selectivity of the sensor was assessed with two different non-complementary PCR products (ds-PCRnon-COM). Diagnostic performance of the biosensor is described and the detection limit is found to be 1.58 ppm. The reliability of the electrochemical biosensing results was verified by electrophoresis of the PCR products.