Isolation and epitope characterization of human monoclonal antibodies to hepatitis C virus core antigen.
In this study we describe the establishment of two hybridoma cell lines secreting human monoclonal antibodies to the 22-kD nucleocapsid protein (core, p22) of the hepatitis C virus (HCV). For this purpose we isolated B lymphocytes from an anti-HCV positive blood donor and infected them with Epstein-Barr (EBV). We obtained several lymphoblastoid cell clones secreting antibodies to the recombinant HCV core protein. The B-cell cultures were oligoclonally expanded and two of them were fused with the (mouse:human) heteromyeloma cell line K6H6/B5. The resulting stable hybridomas produce antibodies of the IgG1/kappa (U1/F10) and the IgM/kappa (Ul/F11) isotype reacting specifically with the recombinant core protein p22. To identify the epitopes recognized by these antibodies we synthesized overlapping peptides (13-mer and 6-mer) from the amino terminus of the core amino acid sequence. Antibody reactivity to these peptides was analyzed in an immunoblot assay. Finally, we were able to define a linear epitope recognized by the Ul/F10 antibody on the nucleocapsid protein. The antibody shows specificity to the sequence N-VYLLPR-C, which corresponds to the amino acids 34-39 of the core sequence.
Siemoneit
K
K
German Red Cross Blood Bank Ulm.
da Silva Cardoso
M
M
Wölpl
A
A
Koerner
K
K
Subanek
B
B
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000956:Antigens, Viral / Q000276:immunology*; D002459:Cell Fusion; D002472:Cell Transformation, Viral; D000939:Epitopes / Q000276:immunology*; D016174:Hepacivirus / Q000276:immunology*; D006508:Hepatitis Antibodies / Q000276:immunology; D004854:Herpesvirus 4, Human; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis; D014758:Viral Core Proteins / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000956:Antigens, Viral; D000939:Epitopes; D006508:Hepatitis Antibodies; D010446:Peptide Fragments; D014758:Viral Core Proteins