HYDRA: Fabrication of cell culture HYDrogels by Robotic liquid handling Automation for high-throughput drug testing

Abstract

Abstract Traditional plastic- and glass-based cell culture platforms lack physiological relevance which limits their predictive power in drug development. Organoids and organs-on-chip offer biomimicry in late preclinical stages but lack scalability for high-throughput screening (HTS) in earlier stages. In fact, even introducing the simplest critical biomimetic cue, thin hydrogel layers, into the small wells of HTS plates causes the formation of curved menisci that disrupt cell seeding and imaging. We introduce HYDRA (HYDrogels by Robotic liquid handling Automation), an automated method that enables consistent hydrogel fabrication in HTS plates. We used liquid handling robots to dispense hydrogel precursor solutions and avoid contact with the sidewalls to prevent meniscus formation. We used gelatin for its mild surfactant properties, which allow pinning of the droplet’s contact line, promoting uniform spread. Finally, we re-aspirated the excess liquid, leaving behind a hydrogel layer with controlled stiffness and thickness. We show compatibility with standard 96- and 384-well plates and integration with existing automated workflows, including quality control. We validated HYDRA in dose-response assays using anti-cancer drugs, genetically engineered epithelial cells, phase holography, and fluorescence microscopy. We believe that HYDRA provides a scalable, biomimetic, and therefore more predictive alternative for early- stage screening and in vitro drug testing.