In vivo Non-Invasive Confocal Fluorescence Imaging Beyond 1700 nm Using Superconducting Nanowire Single-Photon Detectors

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 by extending the excitation and emission wavelengths to the > 1000 nm second near-infrared (NIR-II), also called the short-wavelength infrared (SWIR) window. Here, we show biocompatible core-shell lead sulfide/cadmium sulfide (PbS/CdS) quantum dots emitting at ~1880 nm and superconducting nanowire single photon detectors (SNSPD) for single-photon detection up to 2000 nm, enabling one-photon excitation fluorescence imaging window in the 1700–2000 nm (NIR-IIc) range with 1650 nm excitation, the longest one-photon excitation and emission for in vivo mouse imaging to date. Confocal fluorescence imaging in NIR-IIc reached an imaging depth of ~ 1100 μm through intact mouse head, and enabled non-invasive cellular-resolution imaging in the inguinal lymph nodes (LNs) of mice without any surgery. We achieve In vivo molecular imaging of high endothelial venules (HEVs) with diameter down to ~ 6.6 μm and 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.