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<p>Genetically encoded indicators that can detect concentrations of metabolites and signalling molecules through fluorescence lifetime changes are gaining attention, because they expand the potential for quantitative imaging. These indicators offer advantages over conventional fluorescence intensity-based indicators by minimizing artifacts such as variations in indicator concentration, cellular morphological changes, and focus drift. However, the availability of fluorescence lifetime-based genetically encoded indicators remains limited, particularly those compatible with the widely used conventional 488 nm laser in microscopy. Here, we introduce qMaLioffG, a single green fluorescent protein-based ATP indicator that exhibits a substantial fluorescence lifetime shift (1.1 ns) within physiologically relevant ATP concentrations. This enables quantitative imaging of ATP levels in the cytoplasm and mitochondria under steady-state conditions across various cell types, providing insights into ATP distribution. We demonstrate that qMaLioffG can be used in multicellular systems, applying it to <i>Drosophila</i> brain and HeLa cell spheroids to reveal spatially heterogeneous ATP levels.</p>

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qMaLioffG: a genetically encoded green fluorescence lifetime-based indicator enabling quantitative imaging of intracellular ATP

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年11月13日

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  • Satoshi Arai,
  • Hideki Itoh,
  • Cong Quang Vu,
  • Loan Thi Ngoc Nguyen,
  • Mizuho Nakayama,
  • Masanobu Oshima,
  • Atsuya Morita,
  • Kazuko Okamoto,
  • Satoru Okuda,
  • Aki Teranishi,
  • Madori Osawa,
  • Yoshiteru Tamura,
  • Shigeaki Nonoyama,
  • Megumi Takuma,
  • Toshinori Fujie,
  • Satya Ranjan Sarker,
  • Thankiah Sudhaharan,
  • Akihiro Furube,
  • Tetsuro Katayama,
  • Taketoshi Kiya,
  • E. Birgitte Lane,
  • Tetsuya Kitaguchi

摘要

Genetically encoded indicators that can detect concentrations of metabolites and signalling molecules through fluorescence lifetime changes are gaining attention, because they expand the potential for quantitative imaging. These indicators offer advantages over conventional fluorescence intensity-based indicators by minimizing artifacts such as variations in indicator concentration, cellular morphological changes, and focus drift. However, the availability of fluorescence lifetime-based genetically encoded indicators remains limited, particularly those compatible with the widely used conventional 488 nm laser in microscopy. Here, we introduce qMaLioffG, a single green fluorescent protein-based ATP indicator that exhibits a substantial fluorescence lifetime shift (1.1 ns) within physiologically relevant ATP concentrations. This enables quantitative imaging of ATP levels in the cytoplasm and mitochondria under steady-state conditions across various cell types, providing insights into ATP distribution. We demonstrate that qMaLioffG can be used in multicellular systems, applying it to Drosophila brain and HeLa cell spheroids to reveal spatially heterogeneous ATP levels.