Spheroid‐On‐A‐Drop: A Modular Droplet Microfluidics Platform

Droplet-based microfluidics offers a robust route to fabricate hydrogel microcarriers with high precision and reproducibility. Here, we present Spheroid-on-a-Drop, a pressure-driven microfluidic platform for generating monodisperse gelatin methacryloyl (GelMA) microgels for 3D cell encapsulation and spheroid culture. The device combines a flow-focusing junction for controlled droplet breakup with an upstream spiral microchannel used as a passive hydrodynamic conditioning module to improve suspension uniformity and cell spatial distribution prior to gelation. Operating parameters were systematically optimized to achieve stable droplet generation in the dripping regime and narrow size distributions. The resulting GelMA microgels showed high monodispersity, structural integrity, and tunable swelling behavior across media and temperature conditions; SEM imaging of lyophilized samples indicated a homogeneous internal morphology consistent with uniform crosslinking and high water content. As biological validation, A549 human lung carcinoma cells were encapsulated within GelMA microgels, showing high post-encapsulation viability and the formation of compact 3D spheroids during culture. Overall, Spheroid-on-a-Drop provides a reproducible and tunable method to produce biocompatible GelMA microgels for 3D tumor spheroid modeling and downstream applications in drug screening.