Sequences of hepatitis C virus genotypes and their use as therapeutic and diagnostic agents

• US 20030008274A1
• Filed: 07/06/2001
• Published: 01/09/2003
• Est. Priority Date: 04/27/1993
• Status: Active Grant

• Current Assignee: Innogenetics NV (H.U. Group Holdings, Inc.)
• Original Assignee: Innogenetics NV (H.U. Group Holdings, Inc.)
• Inventors: Maertens, Geert, Stuyver, Lieven
• Granted Patent: US 7,195,765 B2
• Time in Patent Office: Days
• US Class Current: 435/5
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: A61K 39/00 Medicinal preparations containing antigens or antibodies materials for immunoassay G01N33/53
• CPC Class Codes: A61P 1/16 for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
• CPC Class Codes: A61P 31/12 Antivirals
• CPC Class Codes: A61P 31/14 for RNA viruses
• CPC Class Codes: A61P 31/20 for DNA viruses
• CPC Class Codes: C07K 14/005 from viruses
• CPC Class Codes: C12N 2770/24222 New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
• CPC Class Codes: Y02A 50/30 Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Abstract

The present invention relates to a polynucleic acid composition comprising or consisting of at least one polynucleic acid containing 8 or more contiguous nucleotides corresponding to a nucleotide sequence from the region spanning positions 417 to 957 of the Core/E1 region of HCV type 3; and/or the region spanning positions 4664 to 4730 of the NS3 region of HCV type 3; and/or the region spanning positions 4892 to 5292 of the NS3/4 region of HCV type 3; and/or the region spanning positions 8 023 to 8 235 of the NS5 region of the BR36 subgroup of HCV type 3a and/or the coding region of HCV type 4a starting at nucleotide 379 in the core region; and/or the coding region of HCV type 4; and/or the coding region of HCV type 5, with said nucleotide numbering being with respect to the numbering of HCV nucleic acids as shown in Table 1, and with said polynucleic acids containing at least one nucleotide difference with known HCV type 1, and/or HCV type 2 genomes in the above-indicated regions, or the complement thereof.

Claims

1. A composition comprising or consisting of at least one polynucleic acid containing 8 or more contiguous nucleotides selected from at least one of the following HCV sequences:

an HCV type 3 genomic sequence, more particularly in any of the following regions;

the region spanning positions 417 to 957 of the Core/E1 region of HCV subtype 3a,

the region spanning positions 4664 to 4730 of the NS3 region of HCV type 3,

the region spanning positions 4892 to 5292 of the NS3/4 region of HCV type 3,

the region spanning positions 8023 to 8235 of the NS5 region of HCV subtype 3a,

an HCV subtype 3c genomic sequence,

an HCV subtype 2d genomic sequence,

an HCV type 4 genomic sequence,

the coding region of HCV subtype 5a,

with said nucleotide numbering being with respect to the numbering of HCV nucleic acids as shown in Table 1, and with said polynucleic acids containing at least one nucleotide difference with known HCV polynucleic acid sequences in the above-indicated regions, or the complement thereof.

2. A composition according to claim 1, wherein said polynucleic acids correspond to a nucleotide sequence selected from any of the following HCV genomic sequences:

an HCV genomic sequence as having a homology of at least 67%, preferably more than 69%, most preferably 71% or more to any of the sequences as represented in SEQ ID NO 13, 15, 17, 19, 21, 23, 25 or 27 in the region spanning positions 417 to 957 of the Core/E1 region;

an HCV genomic sequence as having a homology of at least 65%, preferably more than 67%, most preferably 69% or more to any of the sequences as represented in SEQ ID NO 19, 21, 23, 25 or 27 in the region spanning positions 574 to 957 of the E1 region;

an HCV genomic sequence, having a homology of at least 79%, more preferably at least 81%, most preferably more than 83% or more to any of the sequences as represented in SEQ ID NO 147 in the region spanning positions 1 to 378 of the Core region,

an HCV genomic sequence having a homology of at least 74%, more preferably at least 76%, most preferably more than 78% or more to any of the sequences as represented in SEQ ID NO 13, 15, 17, 19, 21, 23, 25 or 27 in the region spanning positions 417 to 957 in the Core/E1 region;

an HCV genomic sequence having a homology of at least 74%, preferably more than 76%, most preferably 78% or more to any of the sequences as represented in SEQ ID NO 13, 15, 17, 19, 21, 23, 25 or 27 in the region spanning positions 574 to 957 in the E1 region,

an HCV genomic sequence having a homology of more than 73.5%, preferably more than 74%, most preferably 75% homology to any of the sequence as represented in SEQ ID NO 29 in the region spanning positions 4664 to 4730 of the NS3 region;

an HCV genomic sequence having a homology of more than 70%, preferably more than 72%, most preferably more than 74% homology to any of the sequences as represented in SEQ ID NO 29, 31, 33, 35, 37 or 39 in the region spanning positions 4892 to 5292 in the NS3/NS4 region;

an HCV genomic sequence having a homology of more than 95%, preferably 95,5%, most preferably 96% homology to any of the sequences as represented in SEQ ID NO 5, 7, 1, 3, 9 or 11 in the region spanning positions 8023 to 8235 of the NS5 region;

an HCV genomic sequence of the BR36 subgroup of HCV type 3a having a homology of more than 96%, preferably 96.5%, most preferably 97% homology to any of the sequences as represented in SEQ ID NO 5, 7, 1, 3, 9 or 11 in the region spanning positions 8023 to 8192 of the NS5B region;

an HCV genomic sequence having a homology of more than 79%, more preferably more than 81%, and most preferably more than 83% to the sequence as represented in SEQ ID NO 149 in the region spanning positions 7932 to 8271 in the NS5B region.

3. A composition according to claim 1, wherein said polynucleic acids correspond to a nucleotide sequence selected from any of the following HCV genomic sequences:

an HCV genomic sequence having a homology of more than 85%, preferably more than 86%, most preferably more than 87% homology to any of the sequences as represented in SEQ ID NO 41, 43, 45, 47, 49, 51, 53 or 151 in the region spanning positions 1 to 573 of the Core region;

an HCV genomic sequence having a homology of more than 61%, preferably more than 63%, most preferably more than 65% homology to any of the sequences as represented in SEQ ID NO 41, 43, 45, 47, 49, 51, 53, 153 or 155 in the region spanning positions 574 to 957 of the E1 region;

an HCV genomic sequence having a homology of more than 76.5%, preferably of more than 77%, most preferably of more than 78% homology with any of the sequences as represented in SEQ ID NO 55, 57, 197 or 199 in the region spanning positions 3856 to 4209 of the NS3 region;

an HCV genomic sequence having a homology of more than 68%, preferably of more than 70%, most preferably of more than 72% homology with the sequence as represented in SEQ ID NO 157 in the region spanning positions 980 to 1179 of the E1/E2 region;

an HCV genomic sequence having a homology of more than 57%, preferably more than 59%, most preferably more than 61% homology to any of the sequences as represented in SEQ ID NO 59 or 61 in the region spanning positions 4936 to 5296 of the NS4 region;

an HCV genomic sequence having a homology of more than 93%, preferably more than 93.5%, most preferably more than 94% homology to any of the sequences as represented in SEQ ID NO 159 or 161 in the region spanning positions 7932 to 8271 of the NS5B region.

4. A composition according to claim 1, wherein said polynucleic acids correspond to a nucleotide sequence selected from any of the following HCV genomic sequences:

an HCV genomic sequence having a homology of more than 66%, preferably more than 68%, most preferably more than 70% homology in the E1 region spanning positions 574 to 957 to any of the sequences as represented in SEQ ID NO 118, 120 or 122 in the region spanning positions 1 to 957 of the Core/E1 region;

an HCV genomic sequence having a homology of more than 71%, preferably more than 72%, most preferably more than 74% homology to any of the sequences as represented in SEQ ID NO 118, 120 or 122 in the region spanning positions 379 to 957;

an HCV genomic sequence having a homology of more than 85%, preferably more than 86%, most preferably more than 86.5% homology to any of the sequences as represented in SEQ ID NO 183, 185 or 187 in the region spanning positions 379 to 957 of the E1 region;

an HCV genomic sequence having a homology of more than 81%, preferably more than 83%, most preferably more than 85% homology to the sequence as represented in SEQ ID NO 189 in the region spanning positions 379 to 957 of the E1 region;

an HCV genomic sequence having a homology of more than 85%, preferably more than 87%, most preferably more than 89% homology to any of the sequences as represented in SEQ ID NO 167 or 169 in the region spanning positions 379 to 957 of the E1 region;

an HCV genomic sequence having a homology of more than 79%, preferably more than 81%, most preferably more than 83% homology to any of the sequences as represented in SEQ ID NO 171 or 173 in the region spanning positions 379 to 957 of the E1 region;

an HCV genomic sequence having a homology of more than 84%, preferably more than 86%, most preferably more than 88% homology to the sequence as represented in SEQ ID NO 175 in the region spanning positions 379 to 957 of the E1 region;

an HCV genomic sequence having a homology of more than 83%, preferably more than 85%, most preferably more than 87% homology to the sequence as represented in SEQ ID NO 177 in the region spanning positions 379 to 957 of the E1 region,

an HCV genomic sequence having a homology of more than 76%, preferably more than 78%, most preferably more than 80% homology to the sequence as represented in SEQ ID NO 179 in the region spanning positions 379 to 957 of the E1 region;

an HCV genomic sequence having a homology of more than 84%, preferably more than 86%, most preferably more than 88% homology to the sequence as represented in SEQ ID NO 181 in the region spanning positions 379 to 957 of the E1 region;

an HCV genomic sequence having a homology of more than 73%, preferably more than 75%, most preferably more than 77% homology to any of the sequences as represented in SEQ ID NO 106, 108, 110, 112, 114, or 116 in the region spanning positions 7932 to 8271 of the NS5 region;

an HCV genomic sequence having a homology of more than 88%, preferably more than 89%, most preferably more than 90% homology to any of the sequences as represented in SEQ ID NO 106, 108, 110, or 112 in the region spanning positions 7932 to 8271 of the NS5 region;

an HCV genomic sequence having a homology of more than 88%, preferably more than 89%, most preferably more than 90% homology to any of the sequences as represented in SEQ ID NO 116 or 201 in the region spanning positions 7932 to 8271 of the NS5 region;

an HCV genomic sequence having a homology of more than 87%, preferably more than 89%, most preferably more than 90% homology to the sequence as represented in SEQ ID NO 203 in the region spanning positions 7932 to 8271 of the NS5 region;

an HCV genomic sequence having a homology of more than 85%, preferably more than 87%, most preferably more than 89% homology to the sequence as represented in SEQ ID NO 114 in the region spanning positions 7932 to 8271 of the NS5 region;

an HCV genomic sequence having a homology of more than 86%, preferably more than 87%, most preferably more than 88% homology to the sequence as represented in SEQ ID NO 207 in the region spanning positions 7932 to 8271 of the NS5 region;

an HCV genomic sequence having a homology of more than 84%, preferably more than 86%, most preferably more than 88% homology to the sequence as represented in SEQ ID NO 209 in the region spanning positions 7932 to 8271 of the NS5 region,

an HCV genomic sequence having a homology of more than 81%, preferably more than 83%, most preferably more than 85% homology to the sequence as represented in SEQ ID NO 211 in the region spanning positions 7932 to 8271 of the NS5 region

5. A composition according to claim 1, wherein said polynucleic acids correspond to a nucleotide sequence selected from any of the following HCV genomic sequences

an HCV genomic sequence having a homology of more than 78%, preferably more than 80%, most preferably more than 82% homology to the sequence as represented in SEQ ID NO 143 in the region spanning positions 379 to 957 of the Core/E1 region,

an HCV genomic sequence having a homology of more than 74%, preferably more than 76%, most preferably more than 78% homology to the sequence as represented in SEQ ID NO 143 in the region spanning positions 574 to 957;

an HCV genomic sequence having a homology of more than 87%, preferably more than 89%, most preferably more than 91% homology to the sequence as represented in SEQ ID NO 145 in the region spanning positions 7932 to 8271 of the NS5B region

6. A composition according to any of claims 1 to 5, wherein said polynucleic acid is liable to act as a primer for amplifying the nucleic acid of a certain isolate belonging to the genotype from which the primer is derived.

7. A composition according to any of claims 1 to 5, wherein said polynucleic acid is able to act as a hybridization probe for specific detection and/or classification into types of a nucleic acid containing said nucleotide sequence, with said oligonucleotide being possibly labelled or attached to a solid substrate.

8. Use of a composition according to any of claims 1 to 7 for in vitro detecting the presence of one or more HCV genotypes, more particularly for detecting the presence of a nucleic acid of any of the HCV genotypes having a nucleotide sequence as defined in any of claims 1 to 5, present in a biological sample liable to contain them, comprising at least the following steps:

(i) possibly extracting sample nucleic acid,

(ii) possibly amplifying the nucleic acid with at least one of the primers according to claim 6 or any other HCV type 2, HCV type 3, HCV type 4, HCV type 5 or universal HCV primer,

(iii) hybridizing the nucleic acids of the biological sample, possibly under denatured conditions, and with said nucleic acids being possibly labelled during or after amplification, at appropriate conditions with one or more probes according to claim 7, with said probes being preferably attached to a solid substrate,

(iv) washing at appropriate conditions,

(v) detecting the hybrids formed,

(vi) inferring the presence of one or more HCV genotypes present from the observed hybridization pattern.

9. A composition consisting of or comprising at least one peptide or polypeptide containing in its sequence a contiguous sequence of at least 5 amino acids of an HCV polyprotein encoded by any of the polynucleic acids according to any of claims 1 to 5.

10. A composition according to claim 9, wherein said contiguous sequence contains in its sequence at least one of the following amino acid residues:

L7, Q43, M44, S60, R67, Q70, T71, A79, A87, N106, K115, A127, A190, S130, V134, G142, I144, E152, A157, V158, P165, S177 or Y177, I178, V180 or E180 or F182, R184, I186, H187, T189, A190, S191 or G191, Q192 or L192 or I192 or V192 or E192, N193 or H193 or P193, W194 or Y194, H195, A197 or I197 or V197 or T197, V202, 1203 or L203, Q208, A210, V212, F214, T216, R217 or D217 or E217 or V217, H218 or N218, H219 or V219 or L219, L227 or L227, M231 or E231 or Q23 1, T232 or D232 or A232 or K232, Q235 or I235, A237 or T237, I242, I246, S247, S248, V249, S250 or Y250, I251 or V251 or M251 or F251, D252, T254 or V254, L255 or V255, E256 or A256, M258 or F258 or V258, A260 or Q260 or S260, A261, T264 or Y264, M265, I266 or A266, A267, G268 or T268, F271 or M271 or V271, I277, M280 or H280, I284 or A284 or L84, V274, V291, N292 or S292, R293 or I293 or Y293, Q294 or R294, L297 or I297 or Q297, A299 or K299 or Q299, N303 or T303, T308 or L308, T310 or F310 or A310 or D310 or V310, L313, G317 or Q317, L333, S351, A358, A359, A363, S364, A366, T369, L373, F376, Q386, I387, S392, I399, F402, 1403, R405, D454, A461, A463, T464, K484, Q500, E501, S521, K522, H524, N528, S531, S532, V534, F536, F537, M539, I546, C1282, A1283, H1310, V1312, Q1321, P1368, V1372, V1373, K1405, Q1406, S1409, A1424, A1429, C1435, S1436, S1456, H1496, A1504, D1510, D1529, I1543, N1567, D1556, N1567, M1572, Q1579, L1581, S1583, F1585, V1595, E1606 or T1606, M1611, V1612 or L1612, P1630, C1636, P1651, T1656 or I1656, L1663, V1667, V1677, A1681, H1685, E1687, G1689, V1695, A1700, Q1704, Y1705, A1713, A1714 or S1714, M1718, D1719, A1721 or T1721, R1722, A1723 or V1723, H1726 or G1726, E1730, V1732, F1735, I1736, S1737, R1738, T1739, G1740, Q1741, K1742, Q1743, A1744, T1745, L1746, E1747 or K1747, I1749, A1750, T1751 or A1751, V1753, N1755, K1756, A1757, P1758, A1759, H1762, T1763, Y1764, P2645, A2647, K2650, K2653 or L2653, S2664, N2673, F2680, K2681, L2686, H2692, Q2695 or L2695 or 12695, V2712, F2715, V2719 or Q2719, T2722, T2724, S2725, R2726, G2729, Y2735, H2739, I2748, G2746 or I2746, I2748, P2752 or K2752, P2754 or T2754, T2757 or P2757,

with said notation being composed of a letter representing the amino acid residue by its one-letter code, and a number representing the amino acid numbering according to Kato et al., 1990 as shown in Table 1.

11. A composition according to any of claims 9 or 10, wherein said contiguous sequence is selected from any of the following HCV amino acid sequences:

a sequence having a homology of more than 72%, preferably more than 74%, and most preferably more than 77% homology to any of the amino acid sequences as represented in SEQ ID NO 14, 16, 18, 20, 22, 24, 26 or 28 in the region spanning positions 140 to 319 in the Core/E1 region;

a sequence having a homology of more than 70%, preferably more than 72%, and most preferably more than 75% homology to any of the amino acid sequences as represented in SEQ ID NO 14, 16, 18, 20, 22, 24, 26 or 28 in the E1 region spanning positions 192 to 319;

a sequence having a homology of more than 86%, preferably more than 88%, and most preferably more than 90% homology to the amino acid sequences as represented in SEQ ID NO 148 in the region spanning positions 1 to 110 in the Core region;

a sequence having a homology of more than 76%, preferably more than 78%, most preferably more than 80% to any of the amino acid sequences as represented in SEQ ID NO 30, 32, 34, 36, 38 or 40 in the region spanning positions 1646 to 1764 in the NS3/NS4 region;

a sequence having a homology of more than 81.5%, preferably more than 83%, and most preferably more than 86% homology to any of the amino acid sequences as represented in SEQ ID NO 14, 16, 18, 20, 22, 24, 26 or 28 in the E1 region spanning positions 192 to 319;

a sequence having a homology of more than 86%, preferably more than 88%, most preferably more than 90% to the amino acid sequence as represented in SEQ ID NO 150 in the region spanning positions 2645 to 2757 in the NS5B region;

12. A composition according to any of claims 9 or 10, wherein said contiguous sequence is selected from any of the following HCV amino acid sequences:

a sequence having a homology of more than 80%, preferably more than 82%, most preferably more than 84% homology to any of the amino acid sequences as represented in SEQ ID NO 118, 120, and 122 in the region spanning positions 127 to 319,

a sequence having a homology of more than 73%, preferably more than 75%, most preferably more than 78% homology in the E1 region spanning positions 192 to 319 to any of the amino acid sequences as represented in SEQ ID NO 118, 120, and 122, in the region spanning positions 127 to 319,

a sequence having more than 85%, preferably more than 86%, most preferably more than 87% homology to any of the amino acid sequences as represented in SEQ ID NO 118, 120 or 122, in the region spanning positions 192 to 319.

13. A composition according to any of claims 9 or 10, wherein said contiguous sequence is selected from any of the following HCV amino acid sequences:

a sequence having more than 93%, preferably more than 94%, most preferably more than 95% homology in the region spanning Core positions 1 to 191 to any of the amino acid sequences as represented in SEQ ID NO 42, 44, 46, 48, 50, 52, 54, or 152;

a sequence having more than 73%, preferably more than 74%, most preferably more than 76% homology in the region spanning E1 positions 192 to 319 to any of the amino acid sequences as represented in SEQ ID NO 42, 44, 46, 48, 50, 52, 54, 154 or 156;

a sequence spanning positions 1286 to 1403 of the NS3 region, with said sequence being characterized as having more than 90%, preferably more than 91%, most preferably more than 92% homology to any of the amino acid sequences represented in SEQ ID NO 56 to 58;

a sequence spanning positions 1646 to 1764 of the NS3/4 region, with said sequence being characterized as having more than 66%, more particularly 68%, most particularly 70% or more homology to any of the amino acid sequences as represented in SEQ ID NO 60 or

14. A composition according to any of claims 9 to 10, wherein said contiguous sequence is selected from any of the following HCV amino acid sequences:

a sequence having a more than 83%, preferably more than 85%, most preferably more than 87% homology in the region spanning Core positions 1 to 319 to the amino acid sequence as represented in SEQ ID NO 144;

a sequence having a more than 79%, preferably more than 81%, most preferably more than 84% homology in the region spanning E1 positions 192 to 319 to the amino acid sequence as represented in SEQ D NO 144;

a sequence having more than 95%, more particularly 96%, most particularly 97% or more homology to the amino acid sequence as represented in SEQ ID NO 146, in the region spanning positions 2645 to 2757 of the NS5B region.

15. A composition according to any of claims 9 to 14, wherein said sequence is selected from the following peptides:

16. Recombinant vector, particularly for cloning and/or expression, with said recombinant vector comprising a vector sequence, an appropriate prokaryotic, eukaryotic or viral promoter sequence followed by the nucleotide sequences as defined in claims 1 to 5, with said recombinant vector allowing the expression of any one of the HCV type 2 and/or HCV type 3 and/or type 4 and/or type 5 derived polypeptides according to any of claims 9 to 15 in a prokaryotic, or eukaryotic host, or in living mammals when injected as naked DNA, and more particularly a recombinant vector allowing the expression of any of the following HCV type 2, HCV type 3, type 4 or type 5 polypeptides spanning the following amino acid positions:

a polypeptide starting at position 1 and ending at any position in the region between positions 70 and 326, more particularly a polypeptide spanning positions 1 to 70, 1 to 85, positions 1 to 120, positions 1 to 150, positions 1 to 191, positions 1 to 200, for expression of the Core protein, and positions 1 to 263, positions 1 to 326, for expression of the Core and E1 protein;

a polypeptide starting at any position in the region between positions 117 and 192, and ending at any position in the region between positions 263 and 326, more particularly from positions 119 to 326, for expression of E1, or forms that have the putative membrane anchor deleted (positions 264 to 293 plus or minus 8 amino acids);

a polypeptide starting at any position in the region between positions 1556 and 1688, and ending at any position in the region between positions 1739 and 1764, for expression of the NS4 regions, more particularly a polypeptide starting at position 1658 and ending at position 1711 for expression of the NS4a antigen, and more particularly, a polypeptide starting at position 1712 and ending between positions 1743 and 1972, for example 1712-1743, 1712-1764, 1712-1782, 1712-1972, 1712 to 1782 and 1902 to 1972 for expression of the NS4b protein or parts thereof.

17. A composition according to any of claims 9 to 15, wherein said polypeptide is a recombinant polypeptide expressed by means of an expression vector as defined in claim 16.

18. A composition according to any of claims 9 to 15 or 16, for use in a method for immunizing a mammal, preferably humans, against HCV comprising administratering a sufficient amount of the composition possibly accompanied by pharmaceutically acceptable adjuvants, to produce an immune response, more particularly a vaccine composition including HCV type 3 polypeptides derived from the E1, Core, or NS4 region and/or type 4 and/or type 5 and/or type 2 polypeptides.

19. Antibody raised upon immunization with a composition according to any of claims 9 to 15, 17 or 18, by means of a process according to claim 18, with said antibody being reactive with any of the polypeptides as defined in any of claims 9 to 15, 17 or 18.

20. Process for detecting in vitro HCV present in biological sample liable to contain it, comprising at least the following steps:

(i) contacting the biological sample to be analyzed for the presence of HCV antibodies with any of the compositions according to claims 9 to 15, 17 or 18, preferentially in an immobilized form under appropriate conditions which allow the formation of an immune complex, wherein said polypeptide is preferentially in the form of a biotinylated polypeptide and is covalently bound to a solid substrate by means of streptavidin or avidin complexes,

(ii) removing unbound components,

(iii) incubating the immunecomplexes formed with heterologous antibodies, which specifically bind to the antibodies present in the sample to be analyzed, with said heterologous antibodies having conjugated to a detectable label under appropriate conditions,

(iv) detecting the presence of said immunecomplexes visually or by means of densitometry and inferring the HCV serotype(s) present from the observed hybridization pattern.

21. Use of a composition according to any of claims 9 to 15, 17 or 18, for incorporation into a serotyping assay for detecting one or more serological types of HCV present in a biological sample liable to contain it, more particularly for detecting E1 and NS4 antigens or antibodies of the different types to be detected combined in one assay format, comprising at least the following steps:

(i) contacting the biological sample to be analyzed for the presence of HCV antibodies or antigens of one or more serological types, with at least one of the compositions according to claims 9 to 15, 17 or 18 in an immobilized form under appropriate conditions which allow the formation of an immunecomplex, (wherein said polypeptide is preferentially in the form of a biotinylated polypeptide and is covalently bound to a solid substrate by means of streptavidin or avidin complexes),

(ii) removing unbound components,

(iii) incubating the immunecomplexes formed with heterologous antibodies, which specifically bind to the antibodies present in the sample to be analyzed, with said heterologous antibodies having conjugated to a detectable label under appropriate conditions,

(iv) detecting the presence of said immunecomplexes visually or by means of densitometry and inferring the HCV serological types present from the observed binding pattern.

22. A kit for determining the presence of HCV genotypes as defined in any of claims 1 to 5 present in a biological sample liable to contain them, comprising:

possibly at least one primer composition containing any primer selected from those defined in claim 6 or any other HCV type 2 and/or HCV type 3 and/or HCV type 4 and/or HCV type 5, or universal HCV primers,

at least one probe composition according to claim 7, preferably in combination with other polypeptides or peptides from HCV type 1, type 2 or other types of HCV, with said probes being preferentially immobilized on a solid substrate, and more preferentially on one and the same membrane strip,

a buffer or components necessary for producing the buffer enabling hybridization reaction between these probes and the possibly amplified products to be carried out,

a means for detecting the hybrids resulting from the preceding hybriziation,

possibly also including an automated scanning and interpretation device for infering the HCV genotype(s) present in the sample from the observed hybridization pattern.

23. A kit for determining the presence of HCV antibodies according to any of claims 9 to 15, 17 or 18 present in a biological sample liable to contain them, comprising:

at least one polypeptide composition according to any of claims 9 to 15, 17 or 18, with said polypeptides being preferentially immobilized on a solid substrate, and more preferentially on one and the same membrane strip,

a buffer or components necessary for producing the buffer enabling binding reaction between these polypeptides and the antibodies against HCV present in the biological sample,

a means for detecting the immune complexes formed in the preceding binding reaction,

possibly also including an automated scanning and interpretation device for infering the HCV genotype present in the sample from the observed binding pattern.