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In vivo non-invasive confocal fluorescence imaging beyond 1,700 nm using superconducting nanowire single-photon detectors

Nature Nanotechnology volume 17, pages 653–660 (2022)Cite this article

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Abstract

Light scattering by biological tissues sets a limit to the penetration depth of high-resolution optical microscopy imaging of live mammals in vivo. An effective approach to reduce light scattering and increase imaging depth is to extend the excitation and emission wavelengths to the second near-infrared window (NIR-II) at >1,000 nm, also called the short-wavelength infrared window. Here we show biocompatible core–shell lead sulfide/cadmium sulfide quantum dots emitting at ~1,880 nm and superconducting nanowire single-photon detectors for single-photon detection up to 2,000 nm, enabling a one-photon excitation fluorescence imaging window in the 1,700–2,000 nm (NIR-IIc) range with 1,650 nm excitation—the longest one-photon excitation and emission for in vivo mouse imaging so far. Confocal fluorescence imaging in NIR-IIc reached an imaging depth of ~1,100 μm through an intact mouse head, and enabled non-invasive cellular-resolution imaging in the inguinal lymph nodes of mice without any surgery. We achieve in vivo molecular imaging of high endothelial venules with diameters as small as ~6.6 μm, as well as CD169 + macrophages and CD3 + T cells in the lymph nodes, opening the possibility of non-invasive intravital imaging of immune trafficking in lymph nodes at the single-cell/vessel-level longitudinally.

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Fig. 1: Lead sulfide QDs and superconducting nanowire single-photon detectors enabling fluorescence imaging beyond 1,700 nm.
Fig. 2: Fluorescence imaging in NIR-IIb and NIR-IIc windows.
Fig. 3: Non-invasive in vivo confocal microscopy of intact mouse head in NIR-IIc window.
Fig. 4: Non-invasive in vivo NIR-II confocal microscopy of mouse iLNs.

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Data availability

Source data are provided with this paper. All data that support the findings of this study are presented in the main text and the Supplementary Information. Source Data are provided with this paper.

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Acknowledgements

This study was supported by the National Institutes of Health (NIH DP1-NS-105737, H.D.). We thank K. Taylor from JASCO who helped measuring the UV–vis–NIR absorbance spectrum of water using their V-770 Spectrophotometer. F.R. thanks the Fonds de recherche du Québec—Nature et technologies (FRQNT) for funding (F.R.). Single Quantum acknowledges support from the EIC SME Phase 2 project SQP (grant no. 848827 to R.G., J.W.L., A.F. and J.Q-.D.).

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Author notes

  1. These authors contributed equally: Feifei Wang, Fuqiang Ren, Zhuoran Ma.

Authors and Affiliations

  1. Department of Chemistry and Bio-X, Stanford University, Stanford, CA, USA

    Feifei Wang, Fuqiang Ren, Zhuoran Ma, Chun Xu, Ani Baghdasaryan, Jiachen Li & Hongjie Dai

  2. School of Medicine, Stanford University, Stanford, CA, USA

    Liangqiong Qu

  3. Single Quantum B.V., Delft, the Netherlands

    Ronan Gourgues, Johannes W. N. Los, Andreas Fognini & Jessie Qin-Dregely

  4. Department of Imaging Physics, Delft University of Technology, Delft, the Netherlands

    Iman Esmaeil Zadeh

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Contributions

H.D. and F.W. conceived and designed the experiments. H.D. and F.W. designed the optical system. F.W. set up the optical system. F.W., F.R. and Z.M. performed the experiments. F.R. synthesized the PbS/CdS QD. R.G., I.E.Z., J.W.N.L., A.F. and J.Q-.D. developed the SNSPD optimized in a 1,550–2,000 nm window. F.W., F.R., Z.M., L.Q., C.X., A.B., J.L. and H.D. analysed the data. F.W. and H.D. wrote the manuscript. All authors contributed to the general discussion and revision of the manuscript.

Corresponding author

Correspondence to Hongjie Dai.

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The following authors were employed by Single Quantum and may profit financially: R.G., J.W.L., A.F. and J.Q.D.

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Nature Nanotechnology thanks Eva Sevick-Muraca, Wei Zheng and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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Supplementary information

Supplementary Information

Supplementary Notes 1–3, Figs. 1–18 and Tables 1–3.

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Supplementary Video

Animated mouse brain video from non-invasive in vivo NIR-IIc confocal microscopy images of vasculatures in intact mouse head.

Source data

Source Data Fig. 3

Statistical Source Data for Fig. 3e.

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Wang, F., Ren, F., Ma, Z. et al. In vivo non-invasive confocal fluorescence imaging beyond 1,700 nm using superconducting nanowire single-photon detectors. Nat. Nanotechnol. 17, 653–660 (2022). https://doi.org/10.1038/s41565-022-01130-3

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