A quantitative metric for the comparative evaluation of optical clearing protocols for 3D multicellular spheroids

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Diosdi , A , Hirling , D , Kovacs , M , Toth , T , Harmati , M , Koos , K , Buzas , K , Piccinini , F & Horvath , P 2021 , ' A quantitative metric for the comparative evaluation of optical clearing protocols for 3D multicellular spheroids ' , Computational and Structural Biotechnology Journal , vol. 19 , pp. 1233-1243 . https://doi.org/10.1016/j.csbj.2021.01.040

Title: A quantitative metric for the comparative evaluation of optical clearing protocols for 3D multicellular spheroids
Author: Diosdi, Akos; Hirling, Dominik; Kovacs, Maria; Toth, Timea; Harmati, Maria; Koos, Krisztian; Buzas, Krisztina; Piccinini, Filippo; Horvath, Peter
Other contributor: University of Helsinki, Institute for Molecular Medicine Finland
Date: 2021
Language: eng
Number of pages: 11
Belongs to series: Computational and Structural Biotechnology Journal
ISSN: 2001-0370
DOI: https://doi.org/10.1016/j.csbj.2021.01.040
URI: http://hdl.handle.net/10138/335043
Abstract: 3D multicellular spheroids quickly emerged as in vitro models because they represent the in vivo tumor environment better than standard 2D cell cultures. However, with current microscopy technologies, it is difficult to visualize individual cells in the deeper layers of 3D samples mainly because of limited light penetration and scattering. To overcome this problem several optical clearing methods have been proposed but defining the most appropriate clearing approach is an open issue due to the lack of a gold standard metric. Here, we propose a guideline for 3D light microscopy imaging to achieve single-cell resolution. The guideline includes a validation experiment focusing on five optical clearing protocols. We review and compare seven quality metrics which quantitatively characterize the imaging quality of spheroids. As a test environment, we have created and shared a large 3D dataset including approximately hundred fluorescently stained and optically cleared spheroids. Based on the results we introduce the use of a novel quality metric as a promising method to serve as a gold standard, applicable to compare optical clearing protocols, and decide on the most suitable one for a particular experiment. (C) 2021 The Authors. Published by Elsevier B.V. on behalf of Research Network of Computational and Structural Biotechnology.
Subject: Optical tissue clearing
Spheroid
Light-sheet microscopy
Focus metrics
CELL
MICROSCOPY
CLEAR(T)
1182 Biochemistry, cell and molecular biology
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