Diósdi, Ákos and Hirling, Dominik and Kovács, Mária and Tóth, Tímea and Harmati, Mária and Koós, Krisztián and Buzás, Krisztina and Horváth, Péter (2021) A quantitative metric for the comparative evaluation of optical clearing protocols for 3D multicellular spheroids. COMPUTATIONAL AND STRUCTURAL BIOTECHNOLOGY JOURNAL, 19. pp. 1233-1243. ISSN 2001-0370
|
Text
DiosdiCompStrucBiotJ.pdf Available under License Creative Commons Attribution. Download (2MB) | Preview |
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. © 2021 The Authors
| Item Type: | Article |
|---|---|
| Additional Information: | Synthetic and Systems Biology Unit, Biological Research Centre (BRC), Szeged, H-6726, Hungary Doctoral School of Biology, University of Szeged, Szeged, H-6726, Hungary Doctoral School of Computer Science, University of Szeged, Szeged, H-6701, Hungary Department of Immunology, Faculty of Medicine, Faculty of Science and Informatics, University of Szeged, Szeged, H-6720, Hungary IRCCS Istituto Romagnolo per lo Studio dei Tumori (IRST) “Dino Amadori”, Via Piero Maroncelli 40, Meldola, FC I-47014, Italy Institute for Molecular Medicine Finland, University of Helsinki, Helsinki, FI-00014, Finland Single-Cell Technologies Ltd., Szeged, H-6726, Hungary Export Date: 22 April 2021 Correspondence Address: Horvath, P.; Synthetic and Systems Biology Unit, Hungary; email: horvath.peter@brc.hu Funding Agency and Grant Number: LENDULET-BIOMAG Grant [2018-342]; European Regional Development FundsEuropean Commission [GINOP-2.3.2-15-2016-00006, GINOP-2.3.2-15-2016-00026, GINOP-2.3.2-15-2016-00037]; H2020-discovAIR [874656]; H2020 ATTRACT-SpheroidPicker; Chan Zuckerberg Initiative, Seed Networks for the HCA-DVP; Union for International Cancer Control (UICC) [UICC-YY/678329] Funding text: The authors would like to thank Dr. Ji Ming Wang (National Cancer Institute-Frederick, Frederick, MD, USA) for kindly providing us with the 5-8F cell line; Francesco Pampaloni (Goethe University, Frankfurt, Germany) for providing spheroid screening sample holders; Dora Bokor, PharmD (BRC, Szeged, Hungary) for proofreading the manuscript. Funding: A.D., D.H., M.K., T.T., M.H., K.K., B.K., and P.H. acknowledge support from the LENDULET-BIOMAG Grant (2018-342), from the European Regional Development Funds (GINOP-2.3.2-15-2016-00006, GINOP-2.3.2-15-2016-00026, GINOP-2.3.2-15-2016-00037), from the H2020-discovAIR (874656), from the H2020 ATTRACT-SpheroidPicker, and from Chan Zuckerberg Initiative, Seed Networks for the HCA-DVP. FP acknowledges support from the Union for International Cancer Control (UICC) for a UICC Yamagiwa-Yoshida (YY) Memorial International Cancer Study Grant (ref: UICC-YY/678329). |
| Uncontrolled Keywords: | ARTICLE; MICROSCOPY; human; quantitative analysis; case report; clinical article; human cell; cell culture; practice guideline; validation process; gold standard; Statistical tests; Comparative evaluations; Large dataset; multicellular spheroid; multicellular spheroid; Light-sheet microscopy; Spheroid; Optical clearing; Single cell resolution; Quality metrices; Microscopy imaging; light penetration; optical tissue clearing; Focus metrics; Quantitative metric; |
| Subjects: | R Medicine / orvostudomány > R1 Medicine (General) / orvostudomány általában |
| SWORD Depositor: | MTMT SWORD |
| Depositing User: | MTMT SWORD |
| Date Deposited: | 03 Feb 2022 17:31 |
| Last Modified: | 03 Feb 2022 17:31 |
| URI: | http://real.mtak.hu/id/eprint/137216 |
Actions (login required)
 |
Edit Item |
