Dy. Chien et al., Use of a novel hepatitis C virus (HCV) major-epitope chimeric polypeptide for diagnosis of HCV infection, J CLIN MICR, 37(5), 1999, pp. 1393-1397
The genome of hepatitis C virus (HCV) consists of seven functional regions:
the core, El, E2/NS1, NS2, NS3, NS4, and NS5 regions. The U.S. Food and Dr
ug Administration-licensed 2.0G immunoassay for the detection of anti-HCV u
ses proteins from the core, NS3, and NS4 regions (McHutchinson et al,, Hepa
tology 15:19-25, 1992), The 3.0G enzyme-linked immunosorbent assay includes
the protein from the NS5 region (Uyttendaele et al,, Vox Sang. 66:122-129,
1994), The necessity of detecting antibodies to viral envelope proteins (E
l and E2) and to different genotype samples has been demonstrated previousl
y (Chien et al,, Lancet 342:933, 1993; Lok et al,, Hepatology 18:497-502, 1
993), In this study we have attempted to improve the sensitivity of the ant
i-HCV assay by developing a single multiple-epitope fusion antigen (MEFA; M
EFA-6) which incorporates all of the major immunodominant epitopes from the
seven functional regions of the HCV genome. A nucleic acid sequence consis
ting of proteins from the viral core, EI, E2, NS3, NS4, and NS5 regions and
different subtype-specific regions of the NS4 region was constructed, clon
ed, and expressed in yeast. The epitopes present on this antigen can be det
ected by epitope-specific monoclonal and polyclonal antibodies. In a compet
ition assay, the MEFA-6 protein competed with 83 to 96% of genotype-specifi
c antibodies from HCV genotype-specific peptides, This recombinant antigen
was subsequently used to design an anti-HCV chemiluminescent immunoassay. W
e designed our assay using a monoclonal anti-human immunoglobulin G antibod
y bound to the solid phase. Because MEFA-6 is fused with human superoxide d
ismutase (h-SOD),,ve used an anti-human superoxide dismutase, dimethyl acri
dinium ester-labeled monoclonal antibody for detection. Our results indicat
e that MEFA-6 6 exposes all of the major immunogenic epitopes, Its excellen
t sensitivity and specificity for the detection of clinical seroconversion
are demonstrated by this assay.