Automating Tumor Implantation in Zebrafish Larvae for Cancer Research and Medicine

Zebrafish larval injections, including subcutaneous and brain-orthotopic tumor xenografting as well as intravenous drug administration, have become central techniques in zebrafish-based research. These procedures, however, remain tedious, technically challenging, and highly dependent on operator skill, limiting throughput and reproducibility. To address these limitations, we introduce an advanced robotic system that automates microinjections into zebrafish larvae, significantly improving efficiency, precision, and reproducibility over manual injections. Here, we provide a detailed protocol for operating the automated injector, a high-throughput system capable of delivering liquids, particles, or cancer cells at three different anatomical sites with superior accuracy and speed compared to trained zebrafish researchers. We outline the essential steps, including setup, calibrations, injection procedures, and troubleshooting, to ensure optimal performance. Furthermore, we provide representative examples of results from using the robot and discuss its potential to facilitate large-scale larval injections, including for drug screens and precision medicine. By integrating an automated injector into the workflows, researchers can overcome limitations of manual injections and accelerate preclinical drug discovery and personalized treatment selection.