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<p>Conventional DNA biosensors employing "always-on" fluorophores commonly suffer from nonspecific signal activation caused by nuclease-mediated degradation, thereby compromising their detection accuracy. Here, we develop a novel nucleic acid-activatable fluorophore (NAF) that enables precise biomolecule detection and imaging in live cells. NAF achieved a 92-fold fluorescence enhancement via covalent conjugation to DNA, resulting in a remarkably decreased non-specific signal activation in a 10% FBS solution and in non-target cells. We demonstrated the utilization of NAF for live-cell imaging of tumor-associated miR-21 using DNA duplex probes, extracellular ATP detection, as well as targeted photodynamic therapy via membrane-anchored aptamer nanoprobes. These applications highlighted the capacity of NAF to avoid false-positive signals inherent to traditional fluorophore-quencher systems, thereby enhancing the detection accuracy and therapeutic safety. We anticipate that this NAF will offer a novel paradigm for the construction of fluorogenic DNA-based sensors for precise cellular bioanalysis and targeted therapeutics.</p>

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Engineering Nucleic Acid-Activatable Fluorophores for Precise Cellular Imaging

  • 期刊论文

  • 发表日期：

    2026年7月3日

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  • Yining Liu,
  • Siyu Wen,
  • Wen Chen,
  • Hao Tang,
  • Lijuan Tang,
  • Zhenkun Wu

摘要

Conventional DNA biosensors employing "always-on" fluorophores commonly suffer from nonspecific signal activation caused by nuclease-mediated degradation, thereby compromising their detection accuracy. Here, we develop a novel nucleic acid-activatable fluorophore (NAF) that enables precise biomolecule detection and imaging in live cells. NAF achieved a 92-fold fluorescence enhancement via covalent conjugation to DNA, resulting in a remarkably decreased non-specific signal activation in a 10% FBS solution and in non-target cells. We demonstrated the utilization of NAF for live-cell imaging of tumor-associated miR-21 using DNA duplex probes, extracellular ATP detection, as well as targeted photodynamic therapy via membrane-anchored aptamer nanoprobes. These applications highlighted the capacity of NAF to avoid false-positive signals inherent to traditional fluorophore-quencher systems, thereby enhancing the detection accuracy and therapeutic safety. We anticipate that this NAF will offer a novel paradigm for the construction of fluorogenic DNA-based sensors for precise cellular bioanalysis and targeted therapeutics.