| Project/Area Number |
07807056
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Gastroenterology
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| Research Institution | MIE UNIVERSITY |
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Principal Investigator |
KAITO Masahiko MIE UNIVERSITY,Faculty of MEDICINE,ASSISTANT, 医学部, 助手 (70214244)
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| Co-Investigator(Kenkyū-buntansha) |
WATANABE Shozo MIE UNIVERSITY,HEALTH ADMINISTRATION CENTER,PROFESSOR, 保健管理センター, 教授 (20134934)
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| Project Period (FY) |
1995 – 1996
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| Project Status |
Completed (Fiscal Year 1996)
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| Budget Amount *help |
¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1996: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1995: ¥1,400,000 (Direct Cost: ¥1,400,000)
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| Keywords | HEPATITIS C VIRUS / VIRUS PARTICLE / ELECTRON MICROSCOPE / COLLOIDALGOLD / PCR METHOD / MONOCLONALANTIBODY / PLASMA / COREPARTICLE / GPT |
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| Research Abstract |
In 1994, we reported that HCV particles are 55 to 65 nm spherical particles with fine spikelike projections and have a 30 to 35 nm inner core by immunoelectron microscopic study using anti-HCV E1 antibodies. In this study, an immunoelectron microscopy was carried out to determine the distribution of HCV E1 and E2/NS1 antigens on the HCV particle.
HCV particles (subtype 1b, 108 copies/ml) in the sucrose density gradient fractions were examined the reactivity with polyclonal and monoclonal antibodies to HCV E2/NS1 protein by an indirect immunogold electron microscopy. A double staining of HCV E1 and E2/NS1 antigens on HCV particles was also done by different two ways, (1) : HCV particles were initially reacted with a mixture of four monoclonal antibodies to the HCV E1 protein and staphylococcal Protein A-conjugated colloidal gold particles (5nm) and then these were further reacted with rabbit polyclonal antibody to the HCV E2/NS1 protein and goat anti-rabbit IgG colloidal gold particles (
… More
10nm), (2) : the particles were initially reacted with a mixture of six monoclonal antibodied to the HCV E2/NS1 protein and staphylococcal Protein A-conjugated colloidal gold particles (5nm) and then these were reacted with rabbit polyclonal antibody to the HCV E1 protein and goat anti-rabbit IgG colloidal gold particles (10nm). Then, the samples were stained with 2% phosphotungstic acid for observation by electron microscopy. HCV particle 60nm in diameter that exhibited specific gold labeling when reacted with the rabbit polyclonal antibody to the HCV E2/NS1 (RR6) are observed. A mixture of six monoclonal antibodies to the HCV E2/NS1 protein also produced specific labeling to HCV particles. By the double staining, specific gold labelings of both antibodies to E1 and E2/NS1 proteins were also detected on the surface of the HCV particle. This finding indicated that both E1 and E2/NS1 proteins are really structural proteins of the HCV envelope.
Futhermore, we carried out immunoelectronmicroscopic study using anti-HCV core antibodies. we revealed that HCV core particles have icosahedral symmetry and are 30 to 35 nm in diameter. Less
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