Pneumatically controlled microfluidic synthesis of polymeric nanoparticles for mRNA delivery

Efficient and reproducible gene delivery requires nanoscale carriers with well-controlled physicochemical properties, yet conventional bulk synthesis of polymeric nanoparticles (PNPs) often results in broad size distributions, batch variability, and poor scalability. Here, we present a pneumatically driven microfluidic platform that enables precise, tunable, and high-throughput synthesis of polymeric nanoparticles for mRNA delivery. The device integrates integrates a central air-driven channel with three reagent side channels to generate segmented gas-liquid flow, which enhances mixing, prevents clogging, and provides precise control over nanoparticle formation. By modulating airflow, the system produces PH-PEI-mRNA-heparin (PPH) nanoparticles with predictable and uniform sizes while significantly extending chip lifespan. The resulting nanoparticles exhibit low cytotoxicity, stable surface charge, efficient cellular uptake, robust endosomal escape, and transfection efficiency comparable to or exceeding commercial lipid agents. This study establishes a scalable, reproducible, and cost-effective strategy for producing polymeric mRNA nanocomplexes and offers a versatile platform for advancing microfluidic manufacturing of nucleic acid therapeutics.