The present disclosure provides detection methods employing HCV core lipid binding domain and DNA binding domain monoclonal antibodies. In certain embodiments, the lipid binding domain monoclonal antibody recognizes an epitope in amino acids 141 to 161 of HCV core protein.

The present disclosure provides detection methods employing HCV core lipid binding domain and DNA binding domain monoclonal antibodies or antibody fragments. In certain embodiments, the lipid binding domain monoclonal antibody or antibody fragment recognizes an epitope in amino acids 141 to 161 of HCV core protein and the DNA binding domain antibody or antibody fragment recognizes an epitope in amino acids 95-123 (e.g., in amino acids 99-117) of HCV core protein.

Synthetic representative HCV subtypes, including a 1a and 1b genome, dubbed Bole1a and Bole1b, are provided using an inventive method of Bayesian phylogenetic tree analysis, ancestral sequence reconstruction and covariance analysis. Bole1a branches centrally among 390 full-genome sequences used in its design, a carefully curated 143 sequence full-genome dataset, and separate genomic regions including an independent set of 214 E1E2 sequences from a Baltimore cohort. Bole1a is phylogenetically representative of widely circulating strains. Full genome non-synonymous diversity comparison and 9-mer peptide coverage analysis showed that Bole1a is able to provide more coverage (94% and 78% respectively) than any other sequence in the dataset including H77, a traditional reference sequence. Bole1a also provides unsurpassed epitope coverage when compared to all known T cell epitopes.

A polypeptide comprising the contiguous amino acids 1-198 of SEQ ID NO: 2; a polypeptide, which comprises a contiguous amino acid sequence that is at least about 95% identical to the contiguous amino acids 1-198 of SEQ ID NO: 2, an epitope that is immunoreactive with an antibody that specifically binds to the core protein of hepatitis C virus (HCV), and an epitope that is immunoreactive with an antibody that specifically binds to the NS4 region of HCV; a nucleic acid encoding such a polypeptide; a host cell comprising such a nucleic acid; an immunodiagnostic reagent comprising such a polypeptide; a kit comprising such an immunodiagnostic reagent; and a method of determining the presence, amount, or concentration of anti-HCV antibodies in a test sample.

The present invention generally relates to combination immunoassays, reagents and kits for simultaneous detection of HCV antigens and anti-HCV antibodies in a test sample.

A method of preparing extracellularly assembled higher order antigen from a native lower order antigen the method comprising the following steps: (i) contacting lower order antigen with a solution comprising a reducing agent for a time and under 5 conditions sufficient to reduce one or more native cysteines; and (ii) removing or diluting the reducing agent or contacting the reduced lower order antigen with an oxidising agent, to elicit assembly of lower order antigen from (i) into an assembled higher order antigen; wherein at least 10% of the lower order antigen is converted to higher order antigen in step (ii) and whereby the assembled higher order antigen 10 displays at least reduced binding to non-neutralizing antibodies compared to the lower order antigen and retains binding to at least one neutralizing antibody. A method of producing a vaccine composition comprising following the steps of the method and then mixing the assembled higher order antigen with a pharmaceutically or physiologically acceptable diluent, carrier or adjuvant. A composition comprising a 15 higher order extracellularly assembled antigen, wherein the assembled antigen displays at least reduced binding to a non-neutralizing antibody compared to a native control higher order antigen. Use of the assembled higher order antigen to stimulate an immune response or for the detection and/or isolation of an immune cell such as a B-cell specific for the antigen.

Methods of identifying virus antigens are provided. In particular the antigens induce broadly neutralizing antibodies in Hepatitis C virus infections. Compositions for inducing an immune response are identified by the methods described herein.