Development of the Bioluminescent Immunoassay for the Detection of 5-Hydroxymethylcytosine in Dinoflagellate

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  • WU Chun
    BioMedical Research Institute, National Institute of Advanced Industrial Science and Technology (AIST)
  • KURINOMARU Takaaki
    BioMedical Research Institute, National Institute of Advanced Industrial Science and Technology (AIST)

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<p>Cypridina luciferase is a bioluminescent enzyme that has been used as a reporter either in gene expression reporter assays or in immunoassays accompanied by an antibody. To develop a novel bioluminescent assay for the detection of 5-hydroxymethylcytosine, we first conjugated Cypridina luciferase to the antibody against 5-hydroxymethylcytosine. Next, we performed modifications of guanine bases in the genome DNA samples with 4-azidophenylglyoxal and the biotinylation via the azide-Staudinger ligation, which allowed streptavidin to capture and immobilize the genome DNA samples under mild conditions. The detection of 5-hydroxymethylcytosine in the genome DNA samples was performed with the conjugates between Cypridina luciferase and the reduced antibody, which was also confirmed by a surface plasmon resonance assay with the antibody alone. The results obtained from the bioluminescent assay were in good agreement with that of the surface plasmon resonance assay. We succeeded in the detection of 5hmC in the genome DNA samples from the dinoflagellate Pyrocystis Lunula by using this method.</p>

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  • Analytical Sciences

    Analytical Sciences 35 (3), 301-305, 2019-03-10

    社団法人 日本分析化学会

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