Therapeutic extracellular vesicle preparation via sequential electrophoretic enrichment and counterflow microdialysis

Abstract

A microfluidic platform designed to enhance the development of extracellular vesicle (EV)-based therapeutics is presented. The two-chip system combines rapid electrokinetic concentration and purification of EVs together with counterflow buffer exchange to prepare the purified vesicles for pH gradient-driven drug loading. The first chip employs electrophoretic enrichment to capture vesicles at the surface of a nanoporous membrane, with punctuated operation of the electrokinetic device supporting continuous processing of EVs from milliliter-scale volumes of cell culture supernatant. The second chip performs continuous-flow buffer exchange to protonate the vesicles and form a transmembrane pH gradient prior to passive loading of nucleic acid cargo. The modular components are shown to preserve EV bioactivity throughout the full process, using disposable thermoplastic chips that eliminate the need for device cleaning or regeneration between process runs. The technology enables a rapid and convenient workflow for EV-based therapeutic development addressing key limitations associated with established batch processes.