Preparation of a Multi-epitope Fusion Antigen for Hepatitis C Virus Antibodies Immunoassay
Su Qiudong;Guo Minzhuo;Yi Yao;Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention;
Objective Current Hepatitis C virus(HCV) immunoassays used three to six separate recombinant proteins or synthetic peptides with cumbersome preparation of diagnostic antigens and uneven diagnostic efficiency. It is imperative to prepare a HCV antigen with excellent diagnostic efficiency.Methods Prokaryotic expression and chromatographic purification were applied to prepare multi-epitope HCV diagnostic antigens(named H65) covering six immunodominant regions of HCV major epidemic strains(1 b and 2 a) in China. The immunological activity of H65 antigen was preliminarily evaluated by detection of some hepatitis C virus negative and positive sera.Results With Trx as a tag, the soluble H65 was prepared with the concentration 2.991 mg/ml and purity 94.53%. Western blot analysis demonstrated the antigenicity of H65. Based on the data from 50 HCV positive and negative sera, McNemer test suggested that there was no statistical difference between the ‘Gold standard' and H65-based immunoassay with a Kappa 0.75, while there was also no statistical difference between a commercial kit and H65-based immunoassay with a Kappa 0.75, indicating excellent consistency.Conclusion H65 had good immunological activity, which provided a basis for further application of multi-epitope HCV antigen-based HCV-IgG immunoassay.