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1996 Fiscal Year Final Research Report Summary

PURIFICATION OF HEPATITIS C VIRUS PARTICLES AND ANALYSIS OF ITS STRUCTURAL PROTEINS

Research Project

Project/Area Number 06454212
Research Category

Grant-in-Aid for Scientific Research (B)

Allocation TypeSingle-year Grants
Section一般
Research Field Virology
Research InstitutionJICHI MEDICAL SCHOOL

Principal Investigator

OKAMOTO Hiroaki  JICHI MEDICAL SCHOOL.SCHOOL OF MEDICINE,ASSISTANT PROESSOR, 医学部, 講師 (30177092)

Co-Investigator(Kenkyū-buntansha) TOKITA Hajime  JICHI MEDICAL SCHOOL.SCHOOL OF MEDICINE,ASSISTANT, 医学部, 助手 (70254935)
Project Period (FY) 1994 – 1996
Keywordshepatitis C virus / sucrose density gradient centrifugation / electron microscopy / hepatitis C virus core particles / immune electron microscopy
Research Abstract

The largest obstacle to the viral characterization of HCV was the inability to sustain sufficient replication in the culture. HCV existed in two entities in the circulation, one of which had a high density (1.15-1.21g/cm^3) and was associated with gamma-globulins ; the other had a low density (1.06-1.12g/cm^3), appeared to be free virions and paralleled the infectivity titer. A buoyant density of HCV recovered from the circulation was estimated to be 1.09-1.11g/cm^3. Electron microscopy of plasma from infected individuals visualized viral particles with a diameter of 55-60nm. By peeling off the envelope with detergent such as Nonidet P-40 and Tween 80, particles were obtained, which had a diameter of 33 nm (peaked at 1.24-1.25g/cm^3) and an abilty to bind with monoclonal antibodies raised against oligopeptides mimicking the HCV core protein and IgG fractions from sera of persistently infected individuals. Specificity was determined by using rabbit polyclonal antibodies raised against isolate-specific E2/hypervariable region oligopeptides or mouse monoclonal antibodies directed to HCVcore which had been coupled with colloidal gold particles. This direct immunogold electron microscopic study confirmed that HCV virions are 55 to 60 nm diameter spherical particles and have 33nm inner core.
Density heterogeneities of HCV in the circulation due to the binding of low density lipoproteins and immunoglobulins as well as low virus titers hampered the large-scale preparation of free-virions from HCV RNA-positive plasmas, but we could observe tens of HCV particles per grids in average. Accumulated knowledge and techniques were also useful for the physicochemical analysis of a recently identified human hepatitis virus, GB virus C/hepatitis G virus.

  • Research Products

    (12 results)

All Other

All Publications (12 results)

  • [Publications] Kojima M.: "Influence,of antibodies to the hypervariable reglon of E2/NS1 glycoprotein on the selective replication of hepatitis C virus in chimpanzees" Virology. 204. 665-672 (1994)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Nagayama R.: "IgM antibody to a hepatitis C virus core peptide(cp14)for monitoring activity of liver diseare in patients with hepatitis C" J.Med.Virol.42. 311-317 (1994)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Itoh K.: "Cold activation of complement as a marker of hepatitis C viremia in sera from blood donors." Transfus Sci. 16. 283-289 (1995)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Itoh K.: "Cold activation of complement enhansing with the duration of storage in sera from blood donors with hepatitis C virus RNA." Int Hepatol Commun. 5. 89-96 (1996)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Sato K.: "Association of circulating hepatitis G virus with lipoproteins for a lack of binding with antibodiss." Biochem Biophys Res Commun. 229. 719-725 (1996)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Okamoto H.: "Molecular virology of hepatitis C virus." Viral Hepatitis Rev. 3(印刷中). (1997)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Kojima M,et al.: "Influence of antibodies to the hypervariable region of E2/NS1 glycoproteins on the selective replication of hepatitis C virus in chimpanzees." Virology. 204. 665-672 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Nagayama R,et al.: "IgM antibody to a hepatitis C virus core peptide (CP14) for monitoring activity of liver disease in patients with hepatitis C." J Med Virol. 42. 311-317 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Itoh K,et al.: "Cold activation of complement as a marker of hepatitis C viremia in sera from blood donors." Transfus sci. 16. 283-289 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Itoh K,et al.: "Cold activation of complement enhancing with the duration of storage in sera from blood donors with hepatitis C virus RNA." Int Hepatol Commun. 5. 89-96 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Sato K,et al.: "Association of circulating hepatitis G virus with lipoproteins for a lack of binding with antibodies." Biochem Biophys Res Commun. 229. 719-725 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Okamoto H,et al.: "Molecular virology of hepatitis C virus" Viral Hepatitis Rev. (in press). (1997)

    • Description
      「研究成果報告書概要(欧文)」より

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Published: 1999-03-09  

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