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Inter-laboratory automation of the in vitro micronucleus assay using imaging flow cytometry and deep learning

Published version
Peer-reviewed

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Authors

Verma, Jatin R. 
Rees, Benjamin J. 
Harte, Danielle S. G. 
Haxhiraj, Qiellor 

Abstract

Abstract: The in vitro micronucleus assay is a globally significant method for DNA damage quantification used for regulatory compound safety testing in addition to inter-individual monitoring of environmental, lifestyle and occupational factors. However, it relies on time-consuming and user-subjective manual scoring. Here we show that imaging flow cytometry and deep learning image classification represents a capable platform for automated, inter-laboratory operation. Images were captured for the cytokinesis-block micronucleus (CBMN) assay across three laboratories using methyl methanesulphonate (1.25–5.0 μg/mL) and/or carbendazim (0.8–1.6 μg/mL) exposures to TK6 cells. Human-scored image sets were assembled and used to train and test the classification abilities of the “DeepFlow” neural network in both intra- and inter-laboratory contexts. Harnessing image diversity across laboratories yielded a network able to score unseen data from an entirely new laboratory without any user configuration. Image classification accuracies of 98%, 95%, 82% and 85% were achieved for ‘mononucleates’, ‘binucleates’, ‘mononucleates with MN’ and ‘binucleates with MN’, respectively. Successful classifications of ‘trinucleates’ (90%) and ‘tetranucleates’ (88%) in addition to ‘other or unscorable’ phenotypes (96%) were also achieved. Attempts to classify extremely rare, tri- and tetranucleated cells with micronuclei into their own categories were less successful (≤ 57%). Benchmark dose analyses of human or automatically scored micronucleus frequency data yielded quantitation of the same equipotent concentration regardless of scoring method. We conclude that this automated approach offers significant potential to broaden the practical utility of the CBMN method across industry, research and clinical domains. We share our strategy using openly-accessible frameworks.

Description

Keywords

Genotoxicity and Carcinogenicity, Micronucleus test, Genetic toxicology, Compound screening, Machine learning, High throughput, Image analysis.

Journal Title

Archives of Toxicology

Conference Name

Journal ISSN

0340-5761
1432-0738

Volume Title

95

Publisher

Springer Berlin Heidelberg
Sponsorship
Engineering and Physical Sciences Research Council (EP/N013506/1)
Biotechnology and Biological Sciences Research Council (BB/P026818/1)
National Institutes of Health (R35 GM122547)
Life Science Research Network Wales (LSBF/R3-007)