Fabrication of cell culture hydrogels by robotic liquid handling automation for high-throughput drug testing

Traditional plastic and glass culture lacks physiological relevance, undermining predictive power in drug discovery. Organoids and organs-on-chip improve biomimicry but do not scale to high-throughput screening (HTS). Even simple hydrogel coatings in HTS plates suffer from curved menisci that disrupt seeding and imaging. We present HYDRA (HYDrogels by Robotic liquid-handling Automation), an automated method to fabricate thin, planar hydrogel films directly in standard plates. Liquid-handlers dispense sub-contact volumes without wall wetting; immediate re-aspiration pins the contact line, leaving a uniform layer with controlled stiffness and thickness. Using fish gelatin hydrogel, HYDRA produces meniscus-free coatings compatible with routine 96- and 384-well workflows and plate-scale quality control. HYDRA was validated through imaging-based dose-response assays with anticancer compounds, engineered epithelial monolayers, and long-term holographic and fluorescence microscopy. It preserved pharmacological sensitivity while supporting high-content imaging on soft, biomimetic substrates, offering a practical bridge between physiological relevance and HTS scalability for early in-vitro drug testing.