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Colorimetric genotyping using DNA-modified fluorescent nanospheres

Research Project

Project/Area Number 15350047
Research Category

Grant-in-Aid for Scientific Research (B)

Allocation TypeSingle-year Grants
Section一般
Research Field Analytical chemistry
Research InstitutionKumamoto University

Principal Investigator

IHARA Toshihiro  Kumamoto University, Faculty of Engineering, Associate Professor, 工学部, 助教授 (40253489)

Co-Investigator(Kenkyū-buntansha) JYO Akinori  Kumamoto University, Faculty of Engineering, Professor, 工学部, 教授 (40038047)
Project Period (FY) 2003 – 2005
Project Status Completed (Fiscal Year 2005)
Budget Amount *help
¥15,100,000 (Direct Cost: ¥15,100,000)
Fiscal Year 2005: ¥3,300,000 (Direct Cost: ¥3,300,000)
Fiscal Year 2004: ¥4,700,000 (Direct Cost: ¥4,700,000)
Fiscal Year 2003: ¥7,100,000 (Direct Cost: ¥7,100,000)
KeywordsDNA-modified nanosphere / SNP analysis / gene diagnosis / multiplexed analysis / controlled aggregation / immunoassay / allele typing / p53遺伝子 / 光の三原色
Research Abstract

1.Sphere modification by ODNs (oligodeoxynucleotide) and peptides
To avoid side reactions, modifications were carried out in two steps. First sphere surface was thiolated with cystamine. Maleimide group was modified to the end of DNA or peptide using appropriate bifunctional reagent. Both of them were coupled at ambient temperature to obtain molecular-modified nanospheres in high efficiency.
2.Multiplexed multicolor genotyping using selective aggregation of DNA-modified nanospheres
Three kinds of differently colored (RGB, red/green/blue) nanospheres (40 nm diameter) bearing unique ODNs on their surface were prepared for detecting the p53 gene. Each ODN is complementary to the different part in the 45-mer sample, which is a part of a conservative region of the p53 gene containing one of the hot spots. The RGB ternary system gave the aggregates with specific colors corresponding to the added ODN samples, wild type or mutant. In addition, in the presence of both samples, all of the spheres formed aggregates with white emission in consequence of mixing three primary colors of lights. Present technique should allow us to conduct an allele analysis.
3.Multiplexed multicolor immunoassay using selective aggregation of peptide-modified nanospheres
Antigenic Peptides with cationic (KKKKC) or anionic (DDDDC) pentamer tail were immobilized on luminous nanospheres of 40 nm diameter. A mixture of the antigenic peptides of FAK and c-Myc was immobilized to the spheres with red emission, while that of c-Myc and α-catenin was likewise to green spheres. Multiplexed immunoassay was achieved by adding the antibodies to a mixed dispersed solution of these spheres under appropriate conditions. Anti-FAK and anti-α-catenin antibodies formed aggregates with red and green emissions, respectively. On the other hand, the anti-c-Myc antibody formed aggregates emitting a yellow light. This system enabled us to differentiate 3 antibodies in one vessel from the definite differences in aggregate color.

Report

(4 results)
  • 2005 Annual Research Report   Final Research Report Summary
  • 2004 Annual Research Report
  • 2003 Annual Research Report
  • Research Products

    (28 results)

All 2006 2005 2004 2003 Other

All Journal Article (25 results) Publications (3 results)

  • [Journal Article] Colorimetric Multiplexed Immunoassay Using Specific Aggregation of Antigenic Peptide-Modified Luminous Nanoparticles2006

    • Author(s)
      Toshihiro Ihara
    • Journal Title

      Analytica Chimica Acta (印刷中)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2005 Final Research Report Summary
  • [Journal Article] Colorimetric Immunoassay Using Specific Colocalization of Antigen Peptide-Modified Luminous Nanooarticles2006

    • Author(s)
      T.Ihara, Y.Mori, S.Tanaka, A.Jyo
    • Journal Title

      Anal.Chim.Acta (in press)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2005 Final Research Report Summary
  • [Journal Article] Colorimetric Multiplexed Immunoassay Using Specific Aggregation of Antigenic Peptide-Modified Luminous Nanoparticles2006

    • Author(s)
      Toshihiro Ihara
    • Journal Title

      Analytica Chimica Acta (in press)

    • Related Report
      2005 Annual Research Report
  • [Journal Article] Synthesis of the amidite reagent to built bipyridine units into DNA backbone2005

    • Author(s)
      Toshihiro Ihara
    • Journal Title

      Heterocycles 65・2

      Pages: 293-301

    • NAID

      40006603448

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2005 Final Research Report Summary 2004 Annual Research Report
  • [Journal Article] Asymmetric Cooperativity in Tandem Hybridization of Enantiomeric Metal Complex-Tethered Short Fluorescent DNA Probe2005

    • Author(s)
      Yusuke Kitamura
    • Journal Title

      Chemical Communications

      Pages: 4523-4525

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2005 Annual Research Report 2005 Final Research Report Summary
  • [Journal Article] DNA-Templated Cooperative Formation of the Luminous Lanthanide Complex and Its Analytical Application of Gene Detection2005

    • Author(s)
      Yusuke Kitamura
    • Journal Title

      Chemistry Letters 34・12

      Pages: 1606-1607

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2005 Final Research Report Summary
  • [Journal Article] Photochemical Ligation between Anthracene-DNA Conjugates and Its Analytical Application to Gene Analysis2005

    • Author(s)
      Toshihiro Ihara
    • Journal Title

      Nucleic Acids Symposium Series 49

      Pages: 41-42

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2005 Annual Research Report 2005 Final Research Report Summary
  • [Journal Article] Asymmetric Cooperativity in Tandem Hybridization of the DNA Conjugates Bearing Chiral Metal Complexes2005

    • Author(s)
      Toshihiro Ihara
    • Journal Title

      Nucleic Acids Symposium Series 49

      Pages: 229-230

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2005 Annual Research Report 2005 Final Research Report Summary
  • [Journal Article] Synthesis of the Amidite Reagent to Built Bipyridine Units into DNABackbone2005

    • Author(s)
      T.Ihara, Y.Shirasaka, Y.Sato, Y.Kitamura, K.Okada, M.Tazaki, A.Jyo
    • Journal Title

      Heterocycles 65

      Pages: 293-301

    • NAID

      40006603448

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2005 Final Research Report Summary
  • [Journal Article] Asymmetric Cooperativity in Tandem Hybridization of Enantiomeric Metal Complex-Tethered Short Fluorescent DNA Probe2005

    • Author(s)
      Y.Kitamura, T.Ihara, K.Okada, Y.Tsujimura, Y.Shirasaka, M.Tazaki, A.Jyo
    • Journal Title

      Chem.Commun.

      Pages: 4523-4525

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2005 Final Research Report Summary
  • [Journal Article] DNA-Templated Cooperative Formation of the Luminous Lanthanide Complex and Its Analytical Application of Gene Detection2005

    • Author(s)
      Y.Kitamura, T.Ihara, Y.Tsujimura, M.Tazaki, A.Jyo
    • Journal Title

      Chem.Lett. 34

      Pages: 1606-1607

    • NAID

      10016891599

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2005 Final Research Report Summary
  • [Journal Article] Asymmetric Cooperativity in Tandem Hybridization of the DNA Conjugates Bearing Chiral Metal Complexes2005

    • Author(s)
      T.Ihara, Y.Kitamura, K.Okada, M.Tazaki, A.Jyo
    • Journal Title

      Nucleic Acids Symp.Ser. 49

      Pages: 229-230

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2005 Final Research Report Summary
  • [Journal Article] Photochemical Ligation between Anthracene-DNA Conjugates and Its Analytical Application to Gene Analysis2005

    • Author(s)
      T.Ihara, M.Mukae, M.Tabara, Y.Kitamura, A.Jyo
    • Journal Title

      Nucleic Acids Symp.Ser. 49

      Pages: 41-42

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2005 Final Research Report Summary
  • [Journal Article] DNA-Templated Cooperative Formation of the Luminous Lanthanide Complex an is Analytical Application of Gene Detection2005

    • Author(s)
      Yusuke Kitamura
    • Journal Title

      Chemistry Letters 34・12

      Pages: 1606-1607

    • Related Report
      2005 Annual Research Report
  • [Journal Article] Photochemical ligation of DNA conjugates through anthracene cyclodimer formation and its fidelity to the template sequences2004

    • Author(s)
      Toshihiro Ihara
    • Journal Title

      Journal of the American Chemical Society 126-29

      Pages: 8880-8881

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2005 Final Research Report Summary
  • [Journal Article] Preparation of DNA-modified nanoparticles and preliminary study for Colorimetric SNP analysis using their selective aggregations2004

    • Author(s)
      Toshihiro Ihara
    • Journal Title

      Nucleic Acids Research 32・12

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2005 Final Research Report Summary 2004 Annual Research Report
  • [Journal Article] Preparation of DNA-Modified Nanoparticles and Preliminary Study for Colorimetric SNP Analysis Using Their Selective Aggregations2004

    • Author(s)
      T.Ihara, S.Tanaka, Y.Chikaura, A.Jyo
    • Journal Title

      Nucleic Acids Res. 32

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2005 Final Research Report Summary
  • [Journal Article] Photochemical Ligation of DNA Conjugates through Anthracene Cyclodimer Formation and Its Fidelity to the Template Sequences2004

    • Author(s)
      T.Ihara, T.Fujii, M.Mukae, Y.Kitamura, A.Jyo
    • Journal Title

      J.Am.Chem.Soc. 126

      Pages: 8880-8881

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2005 Final Research Report Summary
  • [Journal Article] Photochemical ligation of DNA conjugates through anthracene cyclodimer formation and its fidelity to the template sequences2004

    • Author(s)
      Toshihiro Ihara
    • Journal Title

      Journal of the American Chemical Society 126・29

      Pages: 8880-8881

    • Related Report
      2004 Annual Research Report
  • [Journal Article] Preparation of DNA-modified polystyrene beads and their application to colorimetry for gene detection2003

    • Author(s)
      Shojiro Tanaka
    • Journal Title

      Biomolecular Chemistry

      Pages: 350-351

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2005 Final Research Report Summary
  • [Journal Article] Metal ion-directed outside binding of small DNA ligand2003

    • Author(s)
      Toshihiro Ihara
    • Journal Title

      Nucleic Acids Research, Supplement 3

      Pages: 85-86

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2005 Final Research Report Summary
  • [Journal Article] Highly enhanced duplex stability of dipyrido [3,2-a : 2',3'-c] phenazine-modified oligonucleotide conjugate2003

    • Author(s)
      Yusuke Kitamura
    • Journal Title

      Nucleic Acids Research, Supplement 3

      Pages: 95-96

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2005 Final Research Report Summary
  • [Journal Article] Preparation of DNA-Modified Polystyrene Beads and Their Application to Colormetrv for Gene Detection2003

    • Author(s)
      S.Tanaka, T.Ihara, Y.Chikaura, A.Jyo
    • Journal Title

      Biomolecular Chemistry -A Bridge for the Future-(Y.Baba (Ed))(Maruzen, Tokyo)

      Pages: 350-351

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2005 Final Research Report Summary
  • [Journal Article] Highly Enhanced Duplex Stability of Dipyrido [3,2-a : 2',3'-c]phenazine-Modified Oligonucleotide Conjugate2003

    • Author(s)
      Y.Kitamura, T.Ihara, Y.Shirasaka, T.Mitsuru, M.Tazaki, A.Jyo
    • Journal Title

      Nucleic Acids Res., Suppl.

      Pages: 95-96

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2005 Final Research Report Summary
  • [Journal Article] Metal Ion-Directed Outside Binding of Small DNA Ligand2003

    • Author(s)
      T.Ihara, T.Ikegami, T.Fujii, Y.Kitamura, S.Sueda, M.Takagi, N.E.Geacintov, A.Jyo
    • Journal Title

      Nucleic Acids Res., Suppl.

      Pages: 85-86

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2005 Final Research Report Summary
  • [Publications] Shojiro Tanaka: "Preparation of DNA-modified polystyrene beads and their application to colorimetry for gene detection"Biomolecular Chemistry. 350-351 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] Toshihiro Ihara: "Metal ion-directed outside binding of small DNA ligand"Nucleic Acids Research, Supplement. 3. 85-86 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] Yusuke Kitamura: "Highly enhanced duplex stability of dipyrido [3,2-a:2',3'-c] phenazine-modified oligonucleotide conjugate"Nucleic Acids Research, Supplement. 3. 95-96 (2003)

    • Related Report
      2003 Annual Research Report

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Published: 2003-04-01   Modified: 2016-04-21  

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