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Elimination of mycoplasmas contaminating cell cultures.

Research Project

Project/Area Number 04557076
Research Category

Grant-in-Aid for Developmental Scientific Research (B)

Allocation TypeSingle-year Grants
Research Field Morphological basic dentistry
Research InstitutionHokkaido University

Principal Investigator

WATANABE Tsuguo  Hokkaido University School of Dentistry Professor, 歯学部, 教授 (10064362)

Co-Investigator(Kenkyū-buntansha) SHIBATA Ken-ichiro  Hokkaido University School of Dentistry Instructor, 歯学部, 助手 (50145265)
Project Period (FY) 1992 – 1993
Project Status Completed (Fiscal Year 1993)
Budget Amount *help
¥8,900,000 (Direct Cost: ¥8,900,000)
Fiscal Year 1993: ¥2,900,000 (Direct Cost: ¥2,900,000)
Fiscal Year 1992: ¥6,000,000 (Direct Cost: ¥6,000,000)
KeywordsCell culture / Mycoplasma contamination / Elimination of mycoplasmas / Mycoplasma orale / Mycoplasma fermentans / HeLa cell / Incubation temperature / 細胞培養 / マイコプラズマの除去 / 加熱 / 酸ホスファターゼ / タンパク・チロシン・ホスファターゼ / マイコプラズマ除去
Research Abstract

Effects of detergents, trypsin, MnCl_2, heating and incubation temperature on viability of mycoplasmas (Mycoplasma orale and Mycoplasma fermentans) most frequently contaminating cell cultures were examined. Based on the results obtained, experiments were carried out to eliminate mycoplasmas from HeLa cells artificially infected with M.orale. As a result, incubation of the contaminated cells at 40。C was suggested to be most effective of all tested, and then the following procedures were established.
1. The contaminated cells (5x10^4) are suspended in 10 ml of DME medium supplemented with 10% FCS (FCS-DME) and incubated at 40。C for 24 hours with vigorous shaking.
2. The cells are sedimented by centrifugation at 1, 000 rpm for 5 min and washed 3 times with 10 ml of FCS-DME.For each washing, a centrifuge tube (15 ml) is changed for a new one.
3. Repeat steps 1 and 2 at least 3 more times.
In the process of studying a variety of properties of M.fermentans, acid phosphatase, purified from cell membranes of the organism, were found to exhibit protein tyrosine phosphatase activity. As the organism penetrates mammalian cells, the enzyme may disturb the tyrosine phosphorylation-dephosphorylation pathway of the cells.

Report

(3 results)
  • 1993 Annual Research Report   Final Research Report Summary
  • 1992 Annual Research Report
  • Research Products

    (10 results)

All Other

All Publications (10 results)

  • [Publications] 野田 守: "Purification and characterization of an acid phosphatase from Mycoplasma fermentans." Microbiology and Immunology. 38. 103-107 (1994)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] 柴田健一郎: "Acid phosphatase purified from Mycoplasma fermentans has protein tyrosine phosphatase-like activity." Infection and Immunity. 62. 313-315 (1994)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] 渡邊継男: "Effects of manganese on growth of Mycoplasma salivarium and Mycoplasma orale." Journal of Clinical Microbiology. 32(in press). (1994)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Noda, Mamoru: "Purification and characterization of an acid phosphatase from Mycoplasma fermentans." Microbiology and Immunology. 38. 103-107 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Shibata, Ken-ichiro: "Acid phosphatase purified from Mycoplasma fermentans has protein tyrosine phosphatase-like activity." Infection and Immunity. 62. 313-315 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Watanabe, Tsuguo: "Effects of manganese on growth of Mycoplasma salivarium and Mycoplasma orale." Journal of Clinical Microbiology. 32 : (in press). (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] 柴田健一郎: "Mycoplasma fermentansの有するホスファターゼの性状" 第20回日本マイコプラズマ学会記録. 27-29 (1993)

    • Related Report
      1993 Annual Research Report
  • [Publications] 柴田健一郎: "Acid phosphatase purified from Mycoplasma fermentans has protein tyrosine phosphatase-like aetivity" Infection and Immunity. 62. 313-315 (1994)

    • Related Report
      1993 Annual Research Report
  • [Publications] 野田守: "Purification and characterization of an acid phosphatase from Mycoplasma fermentans" Microkiology and Jmmunology. 38. 103-107 (1994)

    • Related Report
      1993 Annual Research Report
  • [Publications] 渡邊継男: "Effects of manganese on growth of Mycoplasma salivarium and Mycoplasma orale" Journal of chinical Microkiology. 32(印刷中). (1994)

    • Related Report
      1993 Annual Research Report

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Published: 1992-04-01   Modified: 2016-04-21  

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