Skip to main content

Thank you for visiting You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

A phosphorescent probe for in vivo imaging in the second near-infrared window


In the second near-infrared spectral window (NIR-II; with wavelengths of 1,000–1,700 nm), in vivo fluorescence imaging can take advantage of reduced tissue autofluorescence and lower light absorption and scattering by tissue. Here, we report the development and in vivo application of a NIR-II phosphorescent probe that has lifetimes of hundreds of microseconds and a Stokes shift of 430 nm. The probe is made of glutathione-capped copper–indium–selenium nanotubes, and in acidic environments (pH 5.5–6.5) switches from displaying fluorescence to phosphorescence. In xenograft models of osteosarcoma and breast cancer, intravenous or intratumoral injections of the probe enabled phosphorescence imaging at signal-to-background ratios, spatial resolutions and sensitivities higher than NIR-II fluorescence imaging with polymer-stabilized copper–indium–sulfide nanorods. Phosphorescence imaging may offer superior imaging performance for a range of biomedical uses.

This is a preview of subscription content, access via your institution

Access options

Buy this article

Prices may be subject to local taxes which are calculated during checkout

Fig. 1: Characterizations of CISe nanotubes.
Fig. 2: Photophysical properties of CISe nanotubes.
Fig. 3: Possible mechanism of the generation of NIR-II phosphorescence.
Fig. 4: In vivo biosafety evaluation of CISe nanotubes.
Fig. 5: Ultrahigh tumour-specific imaging by NIR-II phosphorescence.
Fig. 6: Superior penetration depth and resolution of NIR-II phosphorescence imaging.

Similar content being viewed by others

Data availability

The main data supporting the results of this study are available within the paper and its Supplementary Information. The raw and analysed datasets are available in Figshare at (ref. 29). Source data are provided with this paper.


  1. Antaris, A. L. et al. A small-molecule dye for NIR-II imaging. Nat. Mater. 15, 235–242 (2016).

    Article  CAS  Google Scholar 

  2. Hu, Z. H. et al. First-in-human liver-tumour surgery guided by multispectral fluorescence imaging in the visible and near-infrared-I/II windows. Nat. Biomed. Eng. 4, 259–271 (2020).

    Article  Google Scholar 

  3. Chen, X. H., Chen, Y. X., Xin, H. H., Wan, T. & Ping, Y. Near-infrared optogenetic engineering of photothermal nanoCRISPR for programmable genome editing. Proc. Natl Acad. Sci. USA 117, 2395–2405 (2020).

    Article  CAS  Google Scholar 

  4. Wang, W. et al. Metabolic labeling of peptidoglycan with NIR-II dye enables in vivo imaging of gut microbiota. Angew. Chem. Int. Ed. 59, 2628–2633 (2020).

    Article  CAS  Google Scholar 

  5. Fan, Y. et al. Lifetime-engineered NIR-II nanoparticles unlock multiplexed in vivo imaging. Nat. Nanotechnol. 13, 941–946 (2018).

    Article  CAS  Google Scholar 

  6. Zhang, M. et al. Bright quantum dots emitting at 1,600 nm in the NIR-IIb window for deep tissue fluorescence imaging. Proc. Natl Acad. Sci. USA 115, 6590–6595 (2018).

    Article  CAS  Google Scholar 

  7. Lei, Z. et al. Stable, wavelength-tunable fluorescent dyes in the NIR-II region for in vivo high-contrast bioimaging and multiplexed biosensing. Angew. Chem. Int. Ed. 58, 8166–8171 (2019).

    Article  CAS  Google Scholar 

  8. Shapiro, A. et al. Tuning optical activity of IV–VI colloidal quantum dots in the short-wave infrared (SWIR) spectral regime. Chem. Mater. 28, 6409–6416 (2016).

    Article  CAS  Google Scholar 

  9. McHugh, K. J. et al. Biocompatible semiconductor quantum dots as cancer imaging agents. Adv. Mater. 30, 1706356 (2018).

    Article  Google Scholar 

  10. Zhu, S. J. & Chen, X. Y. Overcoming the colour barrier. Nat. Photonics 13, 515–516 (2019).

    Article  CAS  Google Scholar 

  11. Su, Y. et al. Ultralong room temperature phosphorescence from amorphous organic materials toward confidential information encryption and decryption. Sci. Adv. 4, eaas9732 (2018).

    Article  Google Scholar 

  12. Zhao, W. J. et al. Boosting the efficiency of organic persistent room-temperature phosphorescence by intramolecular triplet–triplet energy transfer. Nat. Commun. 10, 1595 (2019).

    Article  Google Scholar 

  13. Xu, J. et al. Large-scale synthesis of long crystalline Cu2-xSe nanowire bundles by water-evaporation-induced self assembly and their application in gas sensing. Adv. Funct. Mater. 19, 1759–1766 (2009).

    Article  CAS  Google Scholar 

  14. Xu, J. et al. Large-scale synthesis and phase transformation of CuSe, CuInSe2, and CuInSe2/CuInS2 core/shell nanowire bundles. ACS Nano 4, 1845–1850 (2010).

    Article  CAS  Google Scholar 

  15. Urano, Y. et al. Selective molecular imaging of viable cancer cells with pH-activatable fluorescence probes. Nat. Med. 15, 104–109 (2009).

    Article  CAS  Google Scholar 

  16. Xia, Y. S. et al. Self-assembly of self-limiting monodisperse supraparticles from polydisperse nanoparticles. Nat. Nanotechnol. 6, 580–587 (2011).

    Article  CAS  Google Scholar 

  17. Mou, M. et al. Aggregation-induced emission properties of hydrothermally synthesized Cu–In–S quantum dots. Chem. Commun. 53, 3357–3360 (2017).

    Article  CAS  Google Scholar 

  18. Jara, D. H., Stamplecoskie, K. G. & Kamat, P. V. Two distinct transitions in CuxInS2 quantum dots. Bandgap versus sub-bandgap excitations in copper-deficient structures. J. Phys. Chem. Lett. 7, 1452–1459 (2016).

    Article  CAS  Google Scholar 

  19. Liu, X., Braun, G. B., Qin, M., Ruoslahti, E. & Sugahara, K. N. In vivo cation exchange in quantum dots for tumor-specific imaging. Nat. Commun. 8, 343 (2017).

    Article  Google Scholar 

  20. Yu, M. X. & Zheng, J. Clearance pathways and tumor targeting of imaging nanoparticles. ACS Nano 9, 6655–6674 (2015).

    Article  CAS  Google Scholar 

  21. Zhang, R. R. et al. Beyond the margins: real-time detection of cancer using targeted fluorophores. Nat. Rev. Clin. Oncol. 14, 347–364 (2017).

    Article  CAS  Google Scholar 

  22. Li, J. & Pu, K. Semiconducting polymer nanomaterials as near-infrared photoactivatable protherapeutics for cancer. Acc. Chem. Res. 53, 752–762 (2020).

    Article  CAS  Google Scholar 

  23. Zhang, K. Y. et al. Long-lived emissive probes for time-resolved photoluminescence bioimaging and biosensing. Chem. Rev. 118, 1770–1839 (2018).

    Article  CAS  Google Scholar 

  24. Gu, Y. Y. et al. High-sensitivity imaging of time-domain near-infrared light transducer. Nat. Photonics 13, 525–531 (2019).

    Article  CAS  Google Scholar 

  25. Xu, G. et al. New generation cadmium-free quantum dots for biophotonics and nanomedicine. Chem. Rev. 116, 12234–12327 (2016).

    Article  CAS  Google Scholar 

  26. Heiden, M. G. V., Cantley, L. C. & Thompson, C. B. Understanding the Warburg effect: the metabolic requirements of cell proliferation. Science 324, 1029–1033 (2009).

    Article  Google Scholar 

  27. Wang, Y. G. et al. A nanoparticle-based strategy for the imaging of a broad range of tumours by nonlinear amplification of microenvironment signals. Nat. Mater. 13, 204–212 (2014).

    Article  CAS  Google Scholar 

  28. Zhao, T. et al. A transistor-like pH nanoprobe for tumour detection and image-guided surgery. Nat. Biomed. Eng. 1, 0006 (2017).

    Article  CAS  Google Scholar 

  29. Chang, B. et al. A phosphorescent probe for in vivo imaging in the second near-infrared window. Figshare (2021).

Download references


This work was supported by the National Natural Science Foundation of China (51803161, 51533007 and 61622117), National Key Research and Development Program of China (2017YFA0205200), Office of Science Biological and Environmental Research Program, US Department of Energy (DE-SC0008397) and National Cancer Institute Centers of Cancer Nanotechnology Excellence grant CCNE-TR U54 CA119367.

Author information

Authors and Affiliations



Z.C., B.C. and T.S. conceived of and designed the study, supervised the project and wrote the manuscript. B.C. performed all of the experiments. Z.H. and J.T. assisted with the design of the study and analysed the data. R.L. and H.L. assisted with the biosafety evaluation. D.L. and C.Q. helped with the animal imaging. Y.R. assisted with the in vitro studies. X.S. contributed to structural analysis by TEM. All authors discussed the results and commented on the manuscript.

Corresponding authors

Correspondence to Zhenhua Hu, Taolei Sun or Zhen Cheng.

Ethics declarations

Competing interests

The authors declare no competing interests.

Additional information

Peer review information Nature Biomedical Engineering thanks Zhen Chao Dong and the other, anonymous, reviewer(s) for their contribution to the peer review of this work. Peer reviewer reports are available.

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Supplementary Information

Supplementary Methods, Discussion, Figs. 1–33, Tables 1–3, References and captions for Supplementary Videos 1–4.

Reporting Summary

Video 1

In vivo video-rate NIR-II intensity imaging of a nude mouse bearing 143B tumours, in lateral position at 24 h post-injection of CISe nanotubes.

Video 2

In vivo video-rate NIR-II intensity imaging of a nude mouse bearing 143B tumours, in supine position at 24 h post-injection of CISe nanotubes.

Video 3

In vivo video-rate NIR-II intensity imaging of a nude mouse bearing 143B tumours, in lateral position at 24 h post-injection of PEG-CIS nanorods.

Video 4

In vivo video-rate NIR-II intensity imaging of a nude mouse bearing 143B tumours, in supine position at 24 h post-injection of PEG-CIS nanorods.

Peer Review File

Source data

Source Data Fig. 2

Source data.

Source Data Fig. 3

Source data.

Source Data Fig. 5

Source data.

Source Data Fig. 6

Source data.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Chang, B., Li, D., Ren, Y. et al. A phosphorescent probe for in vivo imaging in the second near-infrared window. Nat. Biomed. Eng 6, 629–639 (2022).

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI:

This article is cited by


Quick links

Nature Briefing: Cancer

Sign up for the Nature Briefing: Cancer newsletter — what matters in cancer research, free to your inbox weekly.

Get what matters in cancer research, free to your inbox weekly. Sign up for Nature Briefing: Cancer