Rapid isolation and quantification of extracellular vesicles from suspension-adapted human embryonic kidney cells...

Exosomes, a subset of extracellular vesicles (EVs, 30-200-nm diameter), serve as biomolecular snapshots of their cell of origin and vehicles for intercellular communication, playing roles in biological processes, including homeostasis maintenance and immune modulation. The large-scale processing of exosomes for use as therapeutic vectors has been proposed, but these applications are limited by impure, low-yield recoveries from cell culture milieu (CCM). Current isolation methods are also limited by tedious and laborious workflows, especially toward an isolation of EVs from CCM for therapeutic applications. Employed is a rapid (<10 min) EV isolation method on a capillary-channeled polymer fiber spin-down tip format. EVs are isolated from the CCM of suspension-adapted human embryonic kidney cells (HEK293), one of the candidate cell lines for commercial EV production. This batch solid-phase extraction technique allows 10¹² EVs to be obtained from only 100-µl aliquots of milieu, processed using a benchtop centrifuge. The tip-isolated EVs were characterized using transmission electron microscopy, multi-angle light scattering, absorbance quantification, an enzyme-linked immunosorbent assay to tetraspanin marker proteins, and a protein purity assay. It is believed that the demonstrated approach has immediate relevance in research and analytical laboratories, with opportunities for production-level scale-up projected.