Application of ethyl cinnamate based optical tissue clearing and expansion microscopy combined with retrograde perfusion for 3D lung imaging

Autor:	Quanchao Sun, Arif ul Maula Khan, Jonathan P. Sleeman, Vincent Heuveline, Norbert Gretz, Anja Schmaus, Cinzia Brenna, Tiziana Picascia
Rok vydání:	2020
Předmět:	0301 basic medicine Pulmonary and Respiratory Medicine Materials science Clinical Biochemistry ethyl cinnamate expansion microscopy retrograde perfusion three-dimensional imaging lung Mice 03 medical and health sciences chemistry.chemical_compound Imaging Three-Dimensional 0302 clinical medicine Lung imaging Microscopy medicine Retrograde perfusion Animals Lung Molecular Biology Tissue clearing business.industry respiratory system Ethyl cinnamate respiratory tract diseases 3. Good health Perfusion 030104 developmental biology medicine.anatomical_structure Three dimensional imaging Microscopy Fluorescence 030228 respiratory system chemistry Cinnamates Nuclear medicine business
Zdroj:	Experimental Lung Research
ISSN:	1521-0499 0190-2148
Popis:	Purpose: 3D imaging of the lung is not a trivial undertaking as during preparation the lung may collapse. Also serial sections and scans followed by 3D reconstruction may lead to artifacts. The present study aims to figure out the best way to perform 3D imaging in lung research. Materials and Methods: We applied an optical tissue clearing (OTC) method, which uses ethyl cinnamate (ECi) as a fast, non-toxic and cheap clearing solvent, for 3D imaging of retrograde perfused lungs by laser confocal fluorescence microscopy and light sheet fluorescence microscopy. We also introduced expansion microscopy (ExM), a cutting-edge technique, in 3D imaging of lungs. We examined and compared the usefulness of these techniques for 3D lung imaging. The ExM protocol was further extended to paraffin-embedded lung metastases blocks. Results: The MHI148-PEI labeled lung vasculature was visualized by retrograde perfusion combined with trachea ligation and ECi based OTC. As compared with trans-cardiac perfusion, the retrograde perfusion results in a better maintenance of lung morphology. 3D structure of alveoli, vascular branches and cilia in lung were revealed by immunofluorescence staining after ExM. 3D distribution of microvasculature and neutrophil cells in 10 years old paraffin-embedded lung metastases were analyzed by ExM. Conclusions: The retrograde perfusion combined with trachea ligation technique could be applied in the lung research in mice. 3D structure of lung vasculature can be visualized by MHI148-PEI perfusion and ECi based OTC in an efficient way. ExM and immunofluorescence staining protocol is highly recommended to perform 3D imaging of fresh fixed lung as well as paraffin-embedded lung blocks.
Databáze:	OpenAIRE
Externí odkaz:	https://explore.openaire.eu/search/publication?articleId=doi_dedup___::5c3cd0330844429055a3f6e42fc1e87d https://doi.org/10.1080/01902148.2020.1829183 Zobrazit plný text záznamu Plný text ve formátu PDF Plný text ve formátu HTML
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