Sandwich Immunoassay For Hepatitis C Virus Non-structural5A Protein Using A Glassy Carbon Electrode Modified With An Au-MoO₃/Chitosan Nanocomposite

ObjectHCV has been shown to have a worldwide distribution, occurringamong people of all ages, genders, races and regions of the world. Thepatients are often undiagnosed and thus they go untreated. So the earlyfound, isolated and treated of the HCV infection was important for limitingof the spread and controlling of the pathogenic condition. Thedetermination of HCV NS5A plays an important role in detection of HCVdiagnosis, progress and prognosis. Due to its low cost, easy operations,rapid detection, high sensitivity, low detection limit, easy tocommercialization and integration, electrochemical immunosensor wasadopted to screen a large number of specimens. The aim of our work is tobuild a high sensitivity and specificity electrochemical immunosensor todetect HCV NS5A protein with nanocomposite. Methods1. Preparation of the Au-MoO₃/CS nanocomposite. MoO₃nanoparticles powders were dissolved in CS solution and ultrasonicprocessed until uniform suspension was obtained. Then, HAuCl4solutionwas dropped in the magnetic stirring suspension. Owe to the reduction ofthe CS, the HAuCl4was reduced to AuNPs on the surface of MoO₃nanoparticles. The Au-MoO₃/CS nanocomposite was then characterized bySEM. This nanocomposite was modified on the GCE and linked with MAby Au-SH bond.2. Due to the reduction of the trisodium citrate, the HAuCl4wasreduced to AuNPs on the surface of MCM-41. Polyclone antibodies andHRP were linked with MCM-41by Au-SH bond. The sandwich type wasused to detect the HCV NS5A antigen. The CV and EIS were employed tocharacterizations of variously modified immunosensor. The performancesof the biosensor were investigated.ResultsThe MoO₃nanoparticles were uniformly suspended in the CS solutionafter the reaction. And uniform size AuNPs was successfully synthesizedon the surface of the MoO₃nanoparticles. The obtained nanocompositewarranted good conductivity, film-forming ability and biocompatibility.The electrochemical immunosensor based on the nanocomposite exhibiteda wide detection range (1ng·mL^-1to50μg·mL^-1). The electrochemical biosensor experiments showed excellent reproducibility, and outstandingstability, RSD4.5%.ConclusionThe electrochemical biosensor showed the properties of shortoperation time, low limit of detection, high selectivity, and excellentstability toward the HCV NS5A determination and was successfully used todetection HCV in real samples.