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Biocompatible carbon dots with low-saturation-intensity and high-photobleaching-resistance for STED nanoscopy imaging of the nucleolus and tunneling nanotubes in living cells

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Abstract

Many kinds of nanoparticles and organic dyes as fluorescent probes have been used in the stimulated emission depletion (STED) nanoscopy. Due to high toxicity, photobleaching and non-water solubility, these fluorescent probes are hard to apply in living cell imaging. Here, we report a new fluorescence carbon dots (FNCDs) with high photoluminescence quantum yield (56%), low toxicity, anti-photobleaching and good water-solubility that suitable for live-cell imaging can be obtained by doping fluorine element. Moreover, the FNCDs can stain the nucleolus and tunneling nanotubes (TNTs) in the living cell. More importantly, for STED nanoscopy imaging, the FNCDs effectively depleted background signals and improved imaging resolution. Furthermore, the lateral resolution of single FNCDs size under the STED nanoscopy is up to 22.1 nm for FNCDs deposited on a glass slide was obtained. And because of their good water dispersibility, the higher resolution of single FNCDs size in the nucleolus of a living cell can be up to 19.7 nm. After the image optimization steps, the fine fluorescence images of TNTs diameter with ca. 75 nm resolution is obtained living cell, yielding a threefold enhancement compared with that in confocal imaging. Additionally, the FNCDs show excellent photobleaching resistance after 1,000 scan cycles in the STED model. All results show that FNCDs have significant potential for application in STED nanoscopy.

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References

  1. Hell, S. W. Far-field optical nanoscopy. Science2007, 316, 1153–1158.

    CAS  Google Scholar 

  2. Yan, W.; Peng, X.; Qi, J.; Gao, J.; Fan, S. P.; Wang, Q.; Qu, J. L.; Niu, H. B. Dynamic fluorescence lifetime imaging based on acousto-optic deflectors. J. Biomed. Opt.2014, 19, 116004.

    Google Scholar 

  3. Egner, A.; Hell, S. W. Fluorescence microscopy with super-resolved optical sections. Trends Cell Biol.2005, 15, 207–215.

    CAS  Google Scholar 

  4. Abbe, E. Beiträge zur theorie des mikroskops und der mikroskopischen Wahrnehmung. Archiv Mikrosk. Anatomie1873, 9, 413–418.

    Google Scholar 

  5. Rust, M. J.; Bates, M.; Zhuang, X. W. Sub-diffraction-limit imaging by stochastic optical reconstruction microscopy (STORM). Nat. Methods2006, 3, 793–796.

    CAS  Google Scholar 

  6. Hell, S. W.; Wichmann, J. Breaking the diffraction resolution limit by stimulated emission: Stimulated-emission-depletion fluorescence microscopy. Opt. Lett.1994, 19, 780–782.

    CAS  Google Scholar 

  7. Betzig, E.; Patterson, G. H.; Sougrat, R.; Lindwasser, O. W.; Olenych, S.; Bonifacino, J. S.; Davidson, M. W.; Lippincott-Schwartz, J.; Hess, H. F. Imaging intracellular fluorescent proteins at nanometer resolution. Science2006, 313, 1642–1645.

    CAS  Google Scholar 

  8. Gustafsson, M. G. L. Nonlinear structured-illumination microscopy: Wide-field fluorescence imaging with theoretically unlimited resolution. Proc. Natl. Acad. Sci. USA2005, 102, 13081–13086.

    CAS  Google Scholar 

  9. Dertinger, T.; Colyer, R.; Iyer, G.; Weiss, S.; Enderlein, J. Fast, background-free, 3D super-resolution optical fluctuation imaging (SOFI). Proc. Natl. Acad. Sci. USA2009, 106, 22287–22292.

    CAS  Google Scholar 

  10. Yang, X. S.; Xie, H.; Alonas, E.; Liu, Y. J.; Chen, X. Z.; Santangelo, P. J.; Ren, Q. S.; Xi, P.; Jin, D. Y. Mirror-enhanced super-resolution microscopy. Light Sci. Appl.2016, 5, e16134.

    CAS  Google Scholar 

  11. Hell, S. W.; Dyba, M.; Jakobs, S. Concepts for nanoscale resolution in fluorescence microscopy. Curr. Opin. Neurobiol.2004, 14, 599–609.

    CAS  Google Scholar 

  12. Hofmann, M.; Eggeling, C.; Jakobs, S.; Hell, S. W. Breaking the diffraction barrier in fluorescence microscopy at low light intensities by using reversibly photoswitchable proteins. Proc. Natl. Acad. Sci. USA2005, 102, 17565–17569.

    CAS  Google Scholar 

  13. Bossi, M.; Fölling, J.; Dyba, M.; Westphal, V.; Hell, S. W. Breaking the diffraction resolution barrier in far-field microscopy by molecular optical bistability. New J. Phys.2006, 8, 275.

    Google Scholar 

  14. Harke, B.; Keller, J.; Ullal, C. K.; Westphal, V.; Schönle, A.; Hell, S. W. Resolution Scaling in STED Microscopy. Opt. Express2008, 16, 4154–1162.

    Google Scholar 

  15. Chen, Z.; Dong, G. P.; Gao, H. W.; Qiu, J. R. Two-/multi-wavelength light excitation effects in optical materials: From fundamentals to applications. Prog. Mater. Sci.2019, 105, 100568.

    Google Scholar 

  16. Lin, Y. H.; Nienhaus, K.; Nienhaus, G. U. Nanoparticle probes for superresolution fluorescence microscopy. ChemNanoMat2018, 4, 253–264.

    CAS  Google Scholar 

  17. Yang, S. T.; Wang, X.; Wang, H. F.; Lu, F. S.; Luo, P. G.; Cao, L.; Meziani, M. J.; Liu, J. H.; Liu, Y. F.; Chen, M. et al. Carbon dots as nontoxic and high-performance fluorescence imaging agents. J. Phys. Chem. C2009, 113, 18110–18114.

    CAS  Google Scholar 

  18. Jin, D. Y.; Xi, P.; Wang, B. M.; Zhang, L.; Enderlein, J.; Van Oijen, A. M. Nanoparticles for super-resolution microscopy and single-molecule tracking. Nat. Methods2018, 15, 415–423.

    CAS  Google Scholar 

  19. Irvine, S. E.; Staudt, T.; Rittweger, E.; Engelhardt, J.; Hell, S. W. Direct light-driven modulation of luminescence from Mn-doped ZnSe quantum dots. Angew. Chem., Int. Ed.2008, 120, 2725–2728.

    Google Scholar 

  20. Liu, Y. J.; Lu, Y. Q.; Yang, X. S.; Zheng, X. L.; Wen, S. H.; Wang, F.; Vidal, X.; Zhao, J. B.; Liu, D. M.; Zhou, Z. G. et al. Amplified stimulated emission in upconversion nanoparticles for super-resolution nanoscopy. Nature2017, 543, 229–233.

    CAS  Google Scholar 

  21. Zhan, Q. Q.; Liu, H. C.; Wang, B. J.; Wu, Q. S.; Pu, R.; Zhou, C.; Huang, B. R.; Peng, X. Y.; Ågren, H.; He, S. L. Achieving high-efficiency emission depletion nanoscopy by employing cross relaxation in upconversion nanoparticles. Nat. Commun.2017, 8, 1058.

    Google Scholar 

  22. Yu, Y.; Feng, C.; Hong, Y. N.; Liu, J. Z.; Chen, S. J.; Ng, K. M.; Luo, K. Q.; Tang, B. Z. Cytophilic fluorescent bioprobes for long-term cell tracking. Adv. Mater.2011, 23, 3298–3302.

    CAS  Google Scholar 

  23. Thomas, J. A. Optical imaging probes for biomolecules: An introductory perspective. Chem. Soc. Rev.2015, 44, 4494–4500.

    CAS  Google Scholar 

  24. Hanne, J.; Falk, H. J.; Görlitz, F.; Hoyer, P.; Engelhardt, J.; Sahl, S. J.; Hell, S. W. STED nanoscopy with fluorescent quantum dots. Nat. Commun.2015, 6, 7127.

    CAS  Google Scholar 

  25. Ye, S.; Yan, W.; Zhao, M. J.; Peng, X.; Song, J.; Qu, J. L. Low-saturation-intensity, high-photostability, and high-resolution STED nanoscopy assisted by CsPbBr3 quantum dots. Adv. Mater.2018, 30, 1800167.

    Google Scholar 

  26. Ye, S.; Zhao, M. J.; Yu, M. S.; Zhu, M.; Yan, W.; Song, J.; Qu, J. L. Mechanistic investigation of upconversion photoluminescence in all-inorganic perovskite CsPbBrI2 nanocrystals. J. Phys. Chem. C2018, 122, 3152–3156.

    CAS  Google Scholar 

  27. Deutsch, Z.; Neeman, L.; Oron, D. Luminescence upconversion in colloidal double quantum dots. Nat. Nanotechnol.2013, 8, 649–653.

    CAS  Google Scholar 

  28. Li, D. Y.; Qin, W.; Xu, B.; Qian, J.; Tang, B. Z. AIE nanoparticles with high stimulated emission depletion efficiency and photobleaching resistance for long-term super-resolution bioimaging. Adv. Mater.2017, 29, 1703643.

    Google Scholar 

  29. Wang, L. W.; Chen, B. L.; Yan, W.; Yang, Z. G.; Peng, X.; Lin, D. Y.; Weng, X. Y.; Ye, T.; Qu, J. L. Resolution improvement in STED super-resolution microscopy at low power using a phasor plot approach. Nanoscale2018, 10, 16252–16260.

    CAS  Google Scholar 

  30. Peng, X. Y.; Huang, B. R.; Pu, R.; Liu, H. C.; Zhang, T.; Widengren, J.; Zhan, Q. Q.; Ågren, H. Fast upconversion super-resolution microscopy with 10 µs per pixel dwell times. Nanoscale2019, 11, 1563–1569.

    CAS  Google Scholar 

  31. Ye, S.; Chen, G. Y.; Shao, W.; Qu, J. L.; Prasad, P. N. Tuning upconversion through a sensitizer/activator-isolated NaYF4 core/shell structure. Nanoscale2015, 7, 3976–3984.

    CAS  Google Scholar 

  32. Lim, S. Y.; Shen, W.; Gao, Z. Q. Carbon quantum dots and their applications. Chem. Soc. Rev.2015, 44, 362–381.

    CAS  Google Scholar 

  33. Li, H.; Huang, J.; Liu, Y.; Lu, F.; Zhong, J.; Wang, Y.; Li, S. M.; Lifshitz, Y.; Lee, S. T.; Kang, Z. H. Enhanced RuBisCO activity and promoted dicotyledons growth with degradable carbon dots. Nano Res.2019, 12, 1585–1593.

    CAS  Google Scholar 

  34. Li, H. T.; Kang, Z. H.; Liu, Y.; Lee, S. T. Carbon nanodots: Synthesis, properties and applications. J. Mater. Chem.2012, 22, 24230–24253.

    CAS  Google Scholar 

  35. Bartelmess, J.; Quinn, S. J.; Giordani, S. Carbon nanomaterials: Multifunctional agents for biomedical fluorescence and Raman imaging. Chem. Soc. Rev.2015, 44, 4672–4698.

    CAS  Google Scholar 

  36. Li, H.; Huang, J.; Lu, F.; Liu, Y.; Song, Y. X.; Sun, Y. H.; Zhong, J.; Huang, H.; Wang, Y.; Li, S. M. et al. Impacts of carbon dots on rice plants: Boosting the growth and improving the disease resistance. ACS Appl. Bio Mater.2018, 1, 663–672.

    CAS  Google Scholar 

  37. Leménager, G.; De Luca, E.; Sun, Y. P.; Pompa, P. P. Super-resolution fluorescence imaging of biocompatible carbon dots. Nanoscale2014, 6, 8617–8623.

    Google Scholar 

  38. Zhu, S. J.; Song, Y. B.; Zhao, X. H.; Shao, J. R.; Zhang, J. H.; Yang, B. The photoluminescence mechanism in carbon dots (graphene quantum dots, carbon nanodots, and polymer dots): Current state and future perspective. Nano Res.2015, 8, 355–381.

    CAS  Google Scholar 

  39. Xu, Q.; Kuang, T. R.; Liu, Y.; Cai, L. L.; Peng, X. F.; Sreenivasan Sreeprasad, T.; Zhao, P.; Yu, Z. Q.; Li, N. Heteroatom-doped carbon dots: Synthesis, characterization, properties, photoluminescence mechanism and biological applications. J. Mater. Chem. B2016, 4, 7204–7219.

    CAS  Google Scholar 

  40. Li, H.; Kong, W. Q.; Liu, J.; Liu, N. Y.; Huang, H.; Liu, Y.; Kang, Z. H. Fluorescent N-doped carbon dots for both cellular imaging and highly-sensitive catechol detection. Carbon2015, 91, 66–75.

    CAS  Google Scholar 

  41. Reddy, A. L. M.; Srivastava, A.; Gowda, S. R.; Gullapalli, H.; Dubey, M.; Ajayan, P. M. Synthesis of nitrogen-doped graphene films for lithium battery application. ACS Nano2010, 4, 6337–6342.

    CAS  Google Scholar 

  42. Feng, W.; Long, P.; Feng, Y. Y.; Li, Y. Two-dimensional fluorinated graphene: Synthesis, structures, properties and applications. Adv. Sci.2016, 3, 1500413.

    Google Scholar 

  43. Li, H.; Huang, J.; Song, Y. X.; Zhang, M. L.; Wang, H. B.; Lu, F.; Huang, H.; Liu, Y.; Dai, X.; Gu, Z. L. et al. Degradable carbon dots with broad-spectrum antibacterial activity. ACS Appl. Mater. Interfaces2018, 10, 26936–26946.

    CAS  Google Scholar 

  44. Li, H.; Zhang, M. L.; Song, Y. X.; Wang, H. B.; Liu, C. A.; Fu, Y. J.; Huang, H.; Liu, Y.; Kang, Z. H. Multifunctional carbon dot for lifetime thermal sensing, nucleolus imaging and antialgal activity. J. Mater. Chem. B2018, 6, 5708–5717.

    Google Scholar 

  45. Dubois, M.; Guérin, K.; Pinheiro, J. P.; Fawal, Z.; Masin, F.; Hamwi, A. NMR and EPR studies of room temperature highly fluorinated graphite heat-treated under fluorine atmosphere. Carbon2004, 42, 1931–1940.

    CAS  Google Scholar 

  46. Hamwi, A.; Daoud, M.; Cousseins, J. C. Graphite fluorides prepared at room temperature 1. Synthesis and characterization. Synth. Met.1988, 26, 89–98.

    CAS  Google Scholar 

  47. Sun, C. B.; Feng, Y. Y.; Li, Y.; Qin, C. Q.; Zhang, Q. Q.; Feng, W. Solvothermally exfoliated fluorographene for high-performance lithium primary batteries. Nanoscale2014, 6, 2634–2641.

    CAS  Google Scholar 

  48. Struzzi, C.; Scardamaglia, M.; Reckinger, N.; Colomer, J. F.; Sezen, H.; Amati, M.; Gregoratti, L.; Snyders, R.; Bittencourt, C. Fluorination of suspended graphene. Nano Res.2017, 10, 3151–3163.

    CAS  Google Scholar 

  49. Long, P.; Feng, Y. Y.; Cao, C.; Li, Y.; Han, J. K.; Li, S. W.; Peng, C.; Li, Z. Y.; Feng, W. Self-protective room-temperature phosphorescence of fluorine and nitrogen codoped carbon dots. Adv. Funct. Mater.2018, 28, 1800791.

    Google Scholar 

  50. Dong, Y. Q.; Pang, H. C.; Yang, H. B.; Guo, C. X.; Shao, J. W.; Chi, Y. W.; Li, C. M.; Yu, T. Carbon-based dots co-doped with nitrogen and sulfur for high quantum yield and excitation-independent emission. Angew. Chem., Int. Ed.2013, 52, 7800–7804.

    CAS  Google Scholar 

  51. Yang, L.; Wang, Z. R.; Wang, J.; Jiang, W. H.; Jiang, X. W.; Bai, Z. S.; He, Y. P.; Jiang, J. Q.; Wang, D. K.; Yang, L. Doxorubicin conjugated functionalizable carbon dots for nucleus targeted delivery and enhanced therapeutic efficacy. Nanoscale2016, 8, 6801–6809.

    CAS  Google Scholar 

  52. Song, Z. Q.; Quan, F. Y.; Xu, Y. H.; Liu, M. L.; Cui, L.; Liu, J. Q. Multifunctional N,S Co-doped carbon quantum dots with pH- and thermo-dependent switchable fluorescent properties and highly selective detection of glutathione. Carbon2016, 104, 169–178.

    CAS  Google Scholar 

  53. Long, P.; Feng, Y. Y.; Li, Y.; Cao, C.; Li, S. W.; An, H. R.; Qin, C. Q.; Han, J. K.; Feng, W. Solid-state fluorescence of fluorine-modified carbon nanodots aggregates triggered by poly(ethylene glycol). ACS Appl. Mater. Interfaces2017, 9, 37981–37990.

    CAS  Google Scholar 

  54. Wang, Z. F.; Yuan, F. L.; Li, X. H.; Li, Y. C.; Zhong, H. Z.; Fan, L. Z.; Yang, S. H. 53% efficient red emissive carbon quantum dots for high color rendering and stable warm white-light-emitting diodes. Adv. Mater.2017, 29, 1702910.

    Google Scholar 

  55. Yuan, F. L.; Yuan, T.; Sui, L. Z.; Wang, Z. B.; Xi, Z. F.; Li, Y. C.; Li, X. H.; Fan, L. Z.; Tan, Z. A.; Chen, A. M. et al. Engineering triangular carbon quantum dots with unprecedented narrow bandwidth emission for multicolored LEDs. Nat. Commun.2018, 9, 2249.

    Google Scholar 

  56. Jiang, K.; Sun, S.; Zhang, L.; Lu, Y.; Wu, A. G.; Cai, C. Z.; Lin, H. W. Red, green, and blue luminescence by carbon dots: Full-color emission tuning and multicolor cellular imaging. Angew. Chem., Int. Ed.2015, 54, 5360–5363.

    CAS  Google Scholar 

  57. Qu, S. N.; Zhou, D.; Li, D.; Ji, W. Y.; Jing, P. T.; Han, D.; Liu, L.; Zeng, H. B.; Shen, D. Z. Toward efficient orange emissive carbon nanodots through conjugated Sp2 -domain controlling and surface charges engineering. Adv. Mater.2016, 28, 3516–3521.

    CAS  Google Scholar 

  58. Sartori-Rupp, A.; Cordero Cervantes, D.; Pepe, A.; Gousset, K.; Delage, E.; Corroyer-Dulmont, S.; Schmitt, C.; Krijnse-Locker, J.; Zurzolo, C. Correlative cryo-electron microscopy reveals the structure of TNTs in neuronal cells. Nat. Commun.2019, 10, 342.

    Google Scholar 

  59. Rustom, A.; Saffrich, R.; Markovic, I.; Walther, P.; Gerdes, H. H. Nanotubular highways for intercellular organelle transport. Science2004, 303, 1007–1010.

    CAS  Google Scholar 

  60. Astanina, K.; Koch, M.; Jüngst, C.; Zumbusch, A.; Kiemer, A. K. Lipid droplets as a novel cargo of tunnelling nanotubes in endothelial cells. Sci. Rep.2015, 5, 11453.

    Google Scholar 

  61. Ariazi, J.; Benowitz, A.; De Biasi, V.; Den Boer, M. L.; Cherqui, S.; Cui, H. F.; Douillet, N.; Eugenin, E. A.; Favre, D.; Goodman, S. et al. Tunneling nanotubes and gap junctions-their role in long-range intercellular communication during development, health, and disease conditions. Front. Mol. Neurosci.2017, 10, 333.

    Google Scholar 

  62. Wang, L. W.; Yan, W.; Li, R. Z.; Weng, X. Y.; Zhang, J.; Yang, Z. G.; Liu, L. W.; Ye, T.; Qu, J. L. Aberration correction for improving the image quality in STED microscopy using the genetic algorithm. Nanophotonics2018, 7, 1971–1980.

    Google Scholar 

  63. Yan, W.; Yang, Y. L.; Tan, Y.; Chen, X.; Li, Y.; Qu, J. L.; Ye, T. Coherent optical adaptive technique improves the spatial resolution of STED microscopy in thick samples. Photonics Res.2017, 5, 176–181.

    Google Scholar 

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Acknowledgements

We thank X. Peng (Shenzhen University) for great assistance in tunneling nanotubes of live cell. This work was partially supported by the National Key R&D Program of China (No. 2018YFC0910600), the National Natural Science Foundation of China (Nos. 61975132, 61775145, 61525503, 61620106016, 61835009, and 81727804), China Postdoctoral Science Foundation (No. 2019M650211), Guangdong Province Key Area R&D Program (No. 2019B110233004), Project of Department of Education of Guangdong Province (No. 2015KGJHZ002/2016KCXTD007), the Shenzhen Basic Research Project (Nos. JCYJ20170412110212234 and JCYJ20170412105003520), and the Natural Science Foundation of Shenzhen University (2019108).

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Correspondence to Wei Yan, Jun Song or Junle Qu.

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Biocompatible carbon dots with low-saturation-intensity and high-photobleaching-resistance for STED nanoscopy imaging of the nucleolus and tunneling nanotubes in living cells

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Li, H., Ye, S., Guo, J. et al. Biocompatible carbon dots with low-saturation-intensity and high-photobleaching-resistance for STED nanoscopy imaging of the nucleolus and tunneling nanotubes in living cells. Nano Res. 12, 3075–3084 (2019). https://doi.org/10.1007/s12274-019-2554-x

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