Evaluation of throughput and yield characteristics of extracellular vesicles on analytical-scale capillary-channeled polymer fiber columns.

Extracellular vesicles (EVs) are lipid-bound nanoparticles secreted by cells that participate in intercellular communication during physiological and pathological processes. Among EV subpopulations, exosomes (30-150 nm) are widely studied for liquid biopsy diagnostics and therapeutic delivery applications. However, existing EV isolation methods often suffer from low throughput, long processing times, and poor scalability. Hydrophobic interaction chromatography (HIC) using polyester (PET) capillary-channeled polymer (C-CP) fiber stationary phases has previously been demonstrated as a rapid (<15 min), low-cost (∼$5 per column) EV purification platform in a microbore column format. This study extends the previously reported microbore-scale platform to an analytical-scale column configuration (2.1 mm i.d. × 250 mm) evaluated across multiple fiber packing densities, which serves as an intermediate scale-up step toward preparative-scale operation. Using urine-derived EVs, dynamic binding capacities approaching ∼1012 particles per column and a 2.3-fold increase in EV capture relative to the microbore format were achieved. EV yields exceeded 65% with process throughputs of up to ∼1011 EVs min-1 alongside ∼96% reduction in co-isolated protein content. Column reproducibility (%RSD = 3.1%) and stable multi-cycle (n = 10) performance further supported platform robustness. The results demonstrate that scaling up the C-CP fiber column format while maintaining efficient transport behavior supports high-yield, high-purity EV isolation at increased throughput, advancing this platform toward practical preparative-scale purification workflows.