Automated In Vitro Wound Healing Assay

Abstract

Wound healing is crucial for the restoration of the epidermal barrier in response to injury. This dynamic process requires an intricate orchestration of cellular migration and proliferation. Our understanding of the intra-cellular signalling mechanisms that initialize, cease, and balance migration and proliferation during wound healing remains incomplete. Wound healing assays are a frequently used approach to examine this complex relationship in vitro. However, existing assays suffer from limited reproducibility, low throughput, and inflexibility in experimental design. We have developed the Automated Wound Healing (AWH) assay, a high-throughput robotics-assisted technique for reproducible wound creation and automated time-lapse image analysis. Wounds are designed as Computer-aided design (CAD) models and recreated in confluent cell layers by the BioAssemblyBot (BAB) 3D-bioprinting platform. The dynamics of migration and proliferation in individual cells are evaluated over the course of wound closure using live-cell fluorescence microscopy and our image processing pipeline. The AWH assay outperforms the standard scratch assay with enhanced consistency in wound geometry. Our ability to create diverse wound shapes in any multi-well plate with the BAB not only allows for multiple experimental conditions to be analyzed in parallel but also offers versatility in the design of wound healing experiments. Our method emerges as a valuable tool for the automated completion and analysis of high-throughput, reproducible, and adaptable in vitro wound healing assays.