*2.8. The Recombinant HCV Proteins*

The recombinant HCV proteins were used as antigens to stimulate T-cell responses in vitro and as sorbents in an enzyme-linked immunosorbent assay (ELISA) to evaluate antibody production. The proteins were combined into four pools: NS3 (protease domain with a sequence of 1027–1229 aa, helicase domain 1230–1658 aa, immunodominant region 1356–1459 aa, genotype 1b); NS4 (1677–1754 aa and mosaic protein containing regions 1691–1710, 1712–1733, 1921–1940 aa from genotypes 1, 2, 3, and 5); NS5A (the full-length protein 1973–2419 aa and fragments 2061–2302 aa, 2212–2313 aa, genotypes 1b and 1a); the NS5B protein lacking C-terminal hydrophobic 21 amino acid residues

(2420–2990 aa, genotype 1b); as a negative control, we used the nucleocapsid (core) protein (1–90 aa). The recombinant proteins were expressed in *E.coli* and purified by chromatography on Ni-NTA-agarose or on glutathione sepharose, as described previously [30,37–39].
