Solid-phase extraction of exosomes from diverse matrices via a polyester capillary-channeled polymer (C-CP) fiber stationary phase in a spin-down tip format

Abstract

Exosomes, a subset of the extracellular vesicle (EV) group of organelles, hold great potential for biomarker detection, therapeutics, disease diagnosis, and personalized medicine applications. The promise and potential of these applications are hindered by the lack of an efficient means of isolation, characterization, and quantitation. Current methods for exosome and EV isolation (including ultracentrifugation, microfiltration, and affinity-based techniques) result in impure recoveries with regard to remnant matrix species (e.g., proteins, genetic material) and are performed on clinically irrelevant time and volume scales. To address these issues, a polyethylene terephthalate (PET) capillary-channeled polymer (C-CP) fiber stationary phase is employed for the solid-phase extraction (SPE) of EVs from various matrices using a micropipette tip-based format. The hydrophobic interaction chromatography (HIC) processing and a spin-down workflow are carried out using a table-top centrifuge. Capture and subsequent elution of intact, biologically active exosomes are verified via electron microscopy and bioassays. The performance of this method was evaluated by capture and elution of exosome standards from buffer solution and three biologically relevant matrices: mock urine, reconstituted non-fat milk, and exosome-depleted fetal bovine serum (FBS). Recoveries were evaluated using UV-Vis absorbance spectrophotometry and ELISA assay. The dynamic binding capacity (50%) for the 1-cm-long (~ 5 μL bed volume) tips was determined using a commercial exosome product, yielding a value of ~ 7 × 1011 particles. The novel C-CP fiber spin-down tip approach holds promise for the isolation of exosomes and other EVs from various matrices with high throughput, low cost, and high efficiency. [Figure not available: see fulltext.].