The rapid and modular assembly of hybrid nanosystems is central to the development of advanced nanomedicines, particularly for solid tumors such as neuroblastoma. While strain-promoted azide–alkyne cycloaddition (SPAAC) has emerged as a powerful tool for catalyst-free, bioorthogonal conjugation, its full potential remains constrained by the need for precise temporal control over fast-reacting systems1.
In this study, we present a custom-designed microfluidic device fabricated from polylactic acid (PLA) using affordable fused filament 3D printing technology2. The chip allows continuous-flow processing with well-defined residence times, enabling highly reproducible nanoassembly workflows. This setup was employed to drive SPAAC-mediated coupling between azide-functionalized mesoporous silica nanoparticles (MCM-41) or azide-functionalized liposomes and various DBCO-bearing entities: gold nanorods, catalase-loaded nanocapsules, and a fluorescently labeled small molecule (DBCO-Fluor 545). These components were selected to span a range of structural and chemical properties, particularly regarding differences in particle rigidity and steric impedance.
Reactions were limited to a 5-minute residence time in both the microfluidic and conventional agitation conditions. Differential centrifugation was used to isolate assembled nanostructures and remove excess reactants. The microfluidic chip consistently enabled efficient and reproducible conjugation across all systems tested, despite the rapid kinetics and low reagent concentrations typically required for biocompatible settings.
Our results demonstrate that the microfluidic system provides a robust and reproducible platform for the rapid generation of hybrid nanoassemblies. Without requiring elevated concentrations or extended reaction times, this strategy enabled the formation of covalently linked nanosystems across all tested configurations. The method offers a scalable and modular route to the fabrication of functional nanomaterials, with potential applications in drug delivery, enzyme immobilization, intracellular trafficking, and targeted cancer therapy.

Bibliography:
1.
Sletten, E. M. & Bertozzi, C. R. Bioorthogonal Chemistry: Fishing for Selectivity in a Sea of Functionality. Angewandte Chemie International Edition 48, 6974–6998 (2009).
2.
Whitesides, G. M. The origins and the future of microfluidics. Nature 442, 368–373 (2006).