Scratching in Style: 3D Printers as Plotters for Automated and Complex Wound Healing Assays

Scratch wound healing assays remain one of the most commonly used 2D migration assays to obtain a broad overview of the migration behavior of cultured cells. They are easy to perform and cost-effective but not standardized in terms of the geometric dimensions of the scratch, e.g., the scratch width and straightness of the line, especially when performed manually. Furthermore, conventional scratch assays only include a single scratch. This leads to the evaluation of a restricted cell population within the culture plates but not of all cells grown in the chamber. There are commercially available scratch-makers that are highly expensive and lack advanced functions such as scratching complex patterns. However, such complex scratch formations would make it possible to assess the entire cell population in a chamber and evaluate the scratchs impact on, for example, migration-associated protein expression. To address this, we developed a novel method utilizing 3D printers as 2D plotters to allow repeatable scratch dimensions for varying pipette tips. This open-source and cost-effective system can be performed with any plate format from any manufacturer under culture hood conditions. The 2D plotter-based method we developed and validated provides highly reproducible and consistent scratch parameters for in vitro migration analyses.