Near-infrared laser scanning confocal microscopy and its application in bioimaging Chaowei Sun 1 • Yalun Wang 1 • Hequn Zhang 1 • Jun Qian 1 Received: 29 June 2017 / Accepted: 26 December 2017 / Published online: 29 December 2017 Ó Springer Science+Business Media, LLC, part of Springer Nature 2017 Abstract Near-infrared (NIR) fluorescence imaging is an important imaging technology in deep-tissue biomedical imaging and related researches, due to the low absorption and scattering of NIR excitation and/or emission in biological tissues. Laser scanning confocal microscopy (LSCM) plays a significant role in the family of fluorescence microscopy. Due to the introduction of pinhole, it can provide images with optical sectioning, high signal-to- noise ratio and better spatial resolution. In this study, in order to combine the advantages of these two techniques, we set up a fluorescence microscopic imaging system, which can be named as NIR-LSCM. The system was based on a commercially available confocal microscope, utilizing a NIR laser for excitation and a NIR sensitive detector for signal collection. In addition, NIR fluorescent nanoparticles (NPs) were prepared, and utilized for fluorescence imaging of the ear and brain of living mice based on the NIR-LSCM system. The structure of blood vessels at certain depth could be visualized clearly, because of the high-resolution and large-depth imaging capability of NIR-LSCM. Keywords Near-infrared (NIR) Á Laser scanning confocal microscopy (LSCM) Á NIR nanoparticles (NPs) Á In vivo Á Bioimaging 1 Introduction Fluorescence imaging in the optical tissue window of 700–900 nm (a commonly used NIR spectral region) can provide larger imaging depth and better signal-to-noise ratio compared to traditional fluorescence bioimaging in visible spectral region, due to lower light scat- tering and absorption (Welsher et al. 2011; Weissleder 2001; Tao et al. 2012; Luo et al. 2011; Kim et al. 2012). In addition, since autofluorescence from biological tissues in NIR & Jun Qian qianjun@zju.edu.cn 1 State Key Laboratory of Modern Optical Instrumentation, Centre for Optical and Electromagnetic Research, Zhejiang University, Hangzhou 310058, China 123 Opt Quant Electron (2018) 50:35 https://doi.org/10.1007/s11082-017-1309-8