High-Throughput Screening Model to Quantify Re-Epithelialization Kinetics

Abstract

In vitro scratch assays provide an attractive method to study the regulation and mechanisms of epithelial cell proliferation and migration that are involved in wound repair. The assay is performed by introducing a scratch into a confluent cell monolayer and monitoring the re-epithelialization of the "wound" by acquiring images over time. Despite improvements to the method over the past years, high-throughput screenings often require the integration of multiple image analysis and data extraction software tools. We developed a high-throughput image-based scratch assay and an automated processing pipeline that employs a mathematical model to describe re-epithelialization kinetics. We implemented a processing pipeline in Galaxy called KREAP that provides an open source web-based platform to enable scientists from diverse backgrounds to perform reproducible and quantitative repair data analyses in less than 45 minutes.

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“…The repair rate parameter derived from the model was used to calculate the average re‐epithelialization rate (cells/min) from technical triplicates. An integrated pipeline for image analysis, single cell enumeration, and kinetic modelling is available online through the Galaxy platform (Fernandez‐Gutierrez et al., 2018) and a detailed protocol of the assay can be found in Nature Protocol Exchange (Fernandez‐Gutierrez et al., 2020). To determine cell displacement, sequential images per treatment were loaded into Fiji (ImageJ), stacked ( z as time), and converted into a hyperstack.…”