Reduction-triggered fluorescent amplification probe for the detection of endogenous RNAs in living human cells

Kazuhiro Furukawa, Hiroshi Abe, Kayo Hibino, Yasushi Sako, Satoshi Tsuneda, Yoshihiro Ito

    Research output: Contribution to journalArticle

    63 Citations (Scopus)

    Abstract

    Oligonucleotide-templated reactions are attracting attention as a method for RNA detection in living cells. Previously, a reduction-triggered fluorescence probe has been reported that is based on azide reduction to switch fluorescence on. In this article, we report a more advanced probe, a reduction-triggered fluorescent amplification probe that is capable of amplifying a target signal. Azidomethyl fluorescein was newly synthesized and introduced into a probe. Azido-masked fluorescein on the probe showed a strong turn-on fluorescence signal upon oligonucleotide-templated Staudinger reduction. The catalytic reaction of the probe offered a turnover number of 50 as fluorescence readout within 4 h. Finally, probes were introduced into human leukemia HL-60 cells by use of streptolysin O pore-forming peptide. We successfully detected and quantitated the 28S rRNA and β-Actin mRNA signal above the background by flow cytometry. In addition, the same RNA targets were imaged by fluorescence microscopy. The data suggest that a reduction-triggered amplification probe may be a powerful tool in analyzing the localization, transcription, or processing of RNA species in living eukaryotic cells.

    Original languageEnglish
    Pages (from-to)1026-1036
    Number of pages11
    JournalBioconjugate Chemistry
    Volume20
    Issue number5
    DOIs
    Publication statusPublished - 2009 May 20

    Fingerprint

    RNA
    Fluorescent Dyes
    Amplification
    Fluorescence
    Cells
    Fluorescein
    Oligonucleotides
    Azides
    HL-60 Cells
    Eukaryotic Cells
    Fluorescence Microscopy
    Actins
    Flow Cytometry
    Leukemia
    Flow cytometry
    Fluorescence microscopy
    Messenger RNA
    Peptides
    Transcription
    Switches

    ASJC Scopus subject areas

    • Biotechnology
    • Bioengineering
    • Organic Chemistry
    • Pharmaceutical Science
    • Biomedical Engineering
    • Pharmacology

    Cite this

    Reduction-triggered fluorescent amplification probe for the detection of endogenous RNAs in living human cells. / Furukawa, Kazuhiro; Abe, Hiroshi; Hibino, Kayo; Sako, Yasushi; Tsuneda, Satoshi; Ito, Yoshihiro.

    In: Bioconjugate Chemistry, Vol. 20, No. 5, 20.05.2009, p. 1026-1036.

    Research output: Contribution to journalArticle

    Furukawa, Kazuhiro ; Abe, Hiroshi ; Hibino, Kayo ; Sako, Yasushi ; Tsuneda, Satoshi ; Ito, Yoshihiro. / Reduction-triggered fluorescent amplification probe for the detection of endogenous RNAs in living human cells. In: Bioconjugate Chemistry. 2009 ; Vol. 20, No. 5. pp. 1026-1036.
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