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      3D NIR-II Molecular Imaging Distinguishes Targeted Organs with High-Performance NIR-II Bioconjugates

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          Abstract

          <p class="first" id="P2">Greatly reduced scattering in the second near-infrared (NIR-II) region (1000–1700 nm) opens up many new exciting avenues of bioimaging research, yet NIR-II fluorescence imaging is mostly implemented by using nontargeted fluorophores or wide-field imaging setups, limiting the signal-to-background ratio and imaging penetration depth due to poor specific binding and out-of-focus signals. A newly developed high-performance NIR-II bioconjugate enables targeted imaging of a specific organ in the living body with high quality. Combined with a home-built NIR-II confocal set-up, the enhanced imaging technique allows 900 µm-deep 3D organ imaging without tissue clearing techniques. Bioconjugation of two hormones to nonoverlapping NIR-II fluorophores facilitates two-color imaging of different receptors, demonstrating unprecedented multicolor live molecular imaging across the NIR-II window. This deep tissue imaging of specific receptors in live animals allows development of noninvasive molecular imaging of multifarious models of normal and neoplastic organs in vivo, beyond the traditional visible to NIR-I range. The developed NIR-II fluorescence microscopy will become a powerful imaging technique for deep tissue imaging without any physical sectioning or clearing treatment of the tissue. </p>

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          Understanding the structure-function relationships at cellular, circuit, and organ-wide scale requires 3D anatomical and phenotypical maps, currently unavailable for many organs across species. At the root of this knowledge gap is the absence of a method that enables whole-organ imaging. Herein, we present techniques for tissue clearing in which whole organs and bodies are rendered macromolecule-permeable and optically transparent, thereby exposing their cellular structure with intact connectivity. We describe PACT (passive clarity technique), a protocol for passive tissue clearing and immunostaining of intact organs; RIMS (refractive index matching solution), a mounting media for imaging thick tissue; and PARS (perfusion-assisted agent release in situ), a method for whole-body clearing and immunolabeling. We show that in rodents PACT, RIMS, and PARS are compatible with endogenous-fluorescence, immunohistochemistry, RNA single-molecule FISH, long-term storage, and microscopy with cellular and subcellular resolution. These methods are applicable for high-resolution, high-content mapping and phenotyping of normal and pathological elements within intact organs and bodies. Copyright © 2014 Elsevier Inc. All rights reserved.
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              Real-time identification of liver cancers by using indocyanine green fluorescent imaging.

              We have often encountered difficulties in identifying small liver cancers during surgery. Fluorescent imaging using indocyanine green (ICG) has the potential to detect liver cancers through the visualization of the disordered biliary excretion of ICG in cancer tissues and noncancerous liver tissues compressed by the tumor. ICG had been intravenously injected for a routine liver function test in 37 patients with hepatocellular carcinoma (HCC) and 12 patients with metastasis of colorectal carcinoma (CRC) before liver resection. Surgical specimens were investigated using a near-infrared light camera system. Among the 49 subjects, the 26 patients examined during the latter period of the study (20 with HCC and 6 with metastasis) underwent ICG-fluorescent imaging of the liver surfaces before resection. ICG-fluorescent imaging identified all of the microscopically confirmed HCCs (n = 63) and CRC metastases (n = 28) in surgical specimens. Among the 63 HCCs, 8 tumors (13%, including 5 early HCCs) were not evident grossly unless observed by ICG-fluorescent imaging. Five false-positive nodules (4 large regenerative nodules and 1 bile duct proliferation) were identified among the fluorescent lesions. Well-differentiated HCCs appeared as uniformly fluorescing lesions with higher lesion-to-liver contrast than that of moderately or poorly differentiated HCCs (162.6 [71.1-218.2] per pixel vs 67.7 [-6.3-211.2] per pixel, P < .001), while CRC metastases were delineated as rim-fluorescing lesions. Fluorescent microscopy confirmed that fluorescence originated in the cytoplasm and pseudoglands of HCC cells and in the noncancerous liver parenchyma surrounding metastases. ICG-fluorescent imaging before resection identified 21 of the 41 HCCs (51%) and all of the 16 metastases that were examined. ICG-fluorescent imaging enables the highly sensitive identification of small and grossly unidentifiable liver cancers in real time, enhancing the accuracy of liver resection and operative staging. (c) 2009 American Cancer Society.
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                Author and article information

                Journal
                Advanced Materials
                Adv. Mater.
                Wiley
                09359648
                March 2018
                March 2018
                February 15 2018
                : 30
                : 13
                : 1705799
                Affiliations
                [1 ]Department of Chemistry; Stanford University; Stanford CA 94305 USA
                [2 ]Program of Reproductive and Stem Cell Biology; Department of Obstetrics and Gynecology; Stanford University School of Medicine; Stanford CA 94305 USA
                [3 ]IVI Foundation; Valencia 46026 Spain
                [4 ]Department of Materials Science & Engineering; South University of Science & Technology of China; Shenzhen 518055 China
                [5 ]Research Center for Advanced Materials and Biotechnology; Research Institute of Tsinghua University in Shenzhen; Shenzhen 518057 China
                Article
                10.1002/adma.201705799
                5931222
                29446156
                7ae9ec4f-18be-4aa2-a3bf-6d312b86d964
                © 2018

                http://doi.wiley.com/10.1002/tdm_license_1.1

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