A palette of bridged bicycle-strengthened fluorophores

doi:10.1038/s41592-025-02693-4

. 2025 Jun;22(6):1276-1287.

doi: 10.1038/s41592-025-02693-4. Epub 2025 May 19.

A palette of bridged bicycle-strengthened fluorophores

Junwei Zhang¹, Kecheng Zhang¹, Kui Wang², Bo Wang^{3

4}, Siyan Zhu^{3

5}, Hongping Qian⁶, Yumiao Ma⁷, Mengling Zhang¹, Tianyan Liu¹, Peng Chen^{8

9}, Yuan Shen³, Yunzhe Fu¹⁰, Shilin Fang², Xinxin Zhang^{2

11}, Peng Zou^{3

5

12}, Wulan Deng^{3

4

13}, Yu Mu^{2

14}, Zhixing Chen^{15

16

17}

Affiliations

¹ College of Future Technology, Institute of Molecular Medicine, National Biomedical Imaging Center, Beijing Key Laboratory of Cardiometabolic Molecular Medicine, State Key Laboratory of Membrane Biology, Peking University, Beijing, China.
² Institute of Neuroscience, State Key Laboratory of Neuroscience, Center for Excellence in Brain Science and Intelligence Technology, Chinese Academy of Sciences, Shanghai, China.
³ Peking-Tsinghua Center for Life Sciences, Academy for Advanced Interdisciplinary Studies, Peking University, Beijing, China.
⁴ Biomedical Pioneering Innovation Center (BIOPIC), Peking University, Beijing, China.
⁵ College of Chemistry and Molecular Engineering, Synthetic and Functional Biomolecules Center, Beijing National Laboratory for Molecular Sciences, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, PKU-IDG/McGovern Institute for Brain Research, Peking University, Beijing, China.
⁶ College of Biological Sciences and Biotechnology, Beijing Forestry University, Beijing, China.
⁷ BSJ Institute, Beijing, China.
⁸ PKU-Nanjing Institute of Translational Medicine, Nanjing, China.
⁹ Genvivo Biotech, Nanjing, China.
¹⁰ Department of Biomedical Engineering, College of Future Technology, Peking University, Beijing, China.
¹¹ ShanghaiTech University, Shanghai, China.
¹² Chinese Institute for Brain Research (CIBR), Beijing, China.
¹³ Beijing Advanced Innovation Center for Genomics (ICG), Peking University, Beijing, China.
¹⁴ University of Chinese Academy of Sciences, Beijing, China.
¹⁵ College of Future Technology, Institute of Molecular Medicine, National Biomedical Imaging Center, Beijing Key Laboratory of Cardiometabolic Molecular Medicine, State Key Laboratory of Membrane Biology, Peking University, Beijing, China. zhixingchen@pku.edu.cn.
¹⁶ Peking-Tsinghua Center for Life Sciences, Academy for Advanced Interdisciplinary Studies, Peking University, Beijing, China. zhixingchen@pku.edu.cn.
¹⁷ PKU-Nanjing Institute of Translational Medicine, Nanjing, China. zhixingchen@pku.edu.cn.

PMID: 40389608
DOI: 10.1038/s41592-025-02693-4

A palette of bridged bicycle-strengthened fluorophores

Junwei Zhang et al. Nat Methods. 2025 Jun.

. 2025 Jun;22(6):1276-1287.

doi: 10.1038/s41592-025-02693-4. Epub 2025 May 19.

Authors

Affiliations

¹ College of Future Technology, Institute of Molecular Medicine, National Biomedical Imaging Center, Beijing Key Laboratory of Cardiometabolic Molecular Medicine, State Key Laboratory of Membrane Biology, Peking University, Beijing, China.
² Institute of Neuroscience, State Key Laboratory of Neuroscience, Center for Excellence in Brain Science and Intelligence Technology, Chinese Academy of Sciences, Shanghai, China.
³ Peking-Tsinghua Center for Life Sciences, Academy for Advanced Interdisciplinary Studies, Peking University, Beijing, China.
⁴ Biomedical Pioneering Innovation Center (BIOPIC), Peking University, Beijing, China.
⁵ College of Chemistry and Molecular Engineering, Synthetic and Functional Biomolecules Center, Beijing National Laboratory for Molecular Sciences, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, PKU-IDG/McGovern Institute for Brain Research, Peking University, Beijing, China.
⁶ College of Biological Sciences and Biotechnology, Beijing Forestry University, Beijing, China.
⁷ BSJ Institute, Beijing, China.
⁸ PKU-Nanjing Institute of Translational Medicine, Nanjing, China.
⁹ Genvivo Biotech, Nanjing, China.
¹⁰ Department of Biomedical Engineering, College of Future Technology, Peking University, Beijing, China.
¹¹ ShanghaiTech University, Shanghai, China.
¹² Chinese Institute for Brain Research (CIBR), Beijing, China.
¹³ Beijing Advanced Innovation Center for Genomics (ICG), Peking University, Beijing, China.
¹⁴ University of Chinese Academy of Sciences, Beijing, China.
¹⁵ College of Future Technology, Institute of Molecular Medicine, National Biomedical Imaging Center, Beijing Key Laboratory of Cardiometabolic Molecular Medicine, State Key Laboratory of Membrane Biology, Peking University, Beijing, China. zhixingchen@pku.edu.cn.
¹⁶ Peking-Tsinghua Center for Life Sciences, Academy for Advanced Interdisciplinary Studies, Peking University, Beijing, China. zhixingchen@pku.edu.cn.
¹⁷ PKU-Nanjing Institute of Translational Medicine, Nanjing, China. zhixingchen@pku.edu.cn.

PMID: 40389608
DOI: 10.1038/s41592-025-02693-4

Abstract

Organic fluorophores are the keystone of advanced biological imaging. The vast chemical space of fluorophores has been extensively explored in search of molecules with ideal properties. However, within the current molecular constraints, there appears to be a trade-off between high brightness, robust photostability, and tunable biochemical properties. Herein we report a general strategy to systematically boost the performance of donor-acceptor-type fluorophores, such as rhodamines, by leveraging SO₂ and O-substituted azabicyclo[3.2.1] octane auxochromes. These bicyclic heterocycles give rise to a collection of 'bridged' dyes (BD) spanning the ultraviolet and visible range with top-notch quantum efficiencies, enhanced water solubility, and tunable cell-permeability. Notably, these azabicyclic fluorophores showed remarkable photostability compared to their tetramethyl or azetidine analogs while being completely resistant to oxidative photoblueing. Functionalized BD dyes are tailored for applications in single-molecule imaging, super-resolution imaging (STED and SIM) in fixed or live mammalian and plant cells, and live zebrafish imaging and chemogenetic voltage imaging.

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Conflict of interest statement

Competing interests: Z.C., J.Z., and P. C. are the inventors of a patent on the bridged-bicycle strengthened fluorophores (CN2023108090626, patent pending), whose value could be affected by this paper. The other authors declare no competing interests.

References

1. Sahl, S. J., Hell, S. W. & Jakobs, S. Fluorescence nanoscopy in cell biology. Nat. Rev. Mol. Cell Biol. 18, 685–701 (2017). - PubMed - DOI
1. Liu, Z., Lavis, LukeD. & Betzig, E. Imaging live-cell dynamics and structure at the single-molecule level. Mol. Cell. 58, 644–659 (2015). - PubMed - DOI
1. Sigal, Y. M., Zhou, R. & Zhuang, X. Visualizing and discovering cellular structures with super-resolution microscopy. Science 361, 880–887 (2018). - PubMed - PMC - DOI
1. Schermelleh, L. et al. Super-resolution microscopy demystified. Nat. Cell Biol. 21, 72–84 (2019). - PubMed - DOI
1. Wang, L., Frei, M. S., Salim, A. & Johnsson, K. Small-molecule fluorescent probes for live-cell super-resolution microscopy. J. Am. Chem. Soc. 141, 2770–2781 (2018). - DOI

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[1] Sahl, S. J., Hell, S. W. & Jakobs, S. Fluorescence nanoscopy in cell biology. Nat. Rev. Mol. Cell Biol. 18, 685–701 (2017). - PubMed - DOI

[2] Sahl, S. J., Hell, S. W. & Jakobs, S. Fluorescence nanoscopy in cell biology. Nat. Rev. Mol. Cell Biol. 18, 685–701 (2017). - PubMed - DOI

[3] Liu, Z., Lavis, LukeD. & Betzig, E. Imaging live-cell dynamics and structure at the single-molecule level. Mol. Cell. 58, 644–659 (2015). - PubMed - DOI

[4] Liu, Z., Lavis, LukeD. & Betzig, E. Imaging live-cell dynamics and structure at the single-molecule level. Mol. Cell. 58, 644–659 (2015). - PubMed - DOI

[5] Sigal, Y. M., Zhou, R. & Zhuang, X. Visualizing and discovering cellular structures with super-resolution microscopy. Science 361, 880–887 (2018). - PubMed - PMC - DOI

[6] Sigal, Y. M., Zhou, R. & Zhuang, X. Visualizing and discovering cellular structures with super-resolution microscopy. Science 361, 880–887 (2018). - PubMed - PMC - DOI

[7] Schermelleh, L. et al. Super-resolution microscopy demystified. Nat. Cell Biol. 21, 72–84 (2019). - PubMed - DOI

[8] Schermelleh, L. et al. Super-resolution microscopy demystified. Nat. Cell Biol. 21, 72–84 (2019). - PubMed - DOI

[9] Wang, L., Frei, M. S., Salim, A. & Johnsson, K. Small-molecule fluorescent probes for live-cell super-resolution microscopy. J. Am. Chem. Soc. 141, 2770–2781 (2018). - DOI

[10] Wang, L., Frei, M. S., Salim, A. & Johnsson, K. Small-molecule fluorescent probes for live-cell super-resolution microscopy. J. Am. Chem. Soc. 141, 2770–2781 (2018). - DOI

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A palette of bridged bicycle-strengthened fluorophores

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A palette of bridged bicycle-strengthened fluorophores

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