Facile, generic capture and on-fiber differentiation of exosomesviaconfocal immunofluorescence microscopy using a capillary-channeled polymer fiber solid-phase extraction tip

Abstract

There is great interest in advancing methodologies for the isolation and characterization of exosomes (30–150 nm, extracellular vesicles (EVs)) for fundamental biochemical research and liquid biopsy applications.

Rapid isolation and quantification of extracellular vesicles from suspension‐adapted human embryonic kidney cells using capillary‐channeled polymer fiber spin‐down tips

“…C-CP fiber micropipette tips were prepared through the previously described process [35][36][37]40]. The fibers were formed via melt-extrusion from bulk polyester (polyethylene terephthalate, PET) in the Clemson University School of Materials Science, having the form of an eight-pronged shape of ∼24 × 38-µm cross section.…”

“…The C-CP tip method allows for the isolation of highly concentrated EV samples in a quantitative and reproducible manner, using minute (100 µl) sample volumes [35][36][37]40]. These qualities are ideal in the case of small population (analytical) sampling of large-scale cell culture conditions to monitor the health of the cell line based on EV production.…”

“…Marcus and colleagues have developed a hydrophobicity‐based EV isolation method employing capillary‐channeled polymer (C‐CP) fiber stationary phases to address the shortcomings of the currently available EV isolation methods [ 32 , 33 , 34 , 35 , 36 , 37 , 38 , 39 , 40 ]. These C‐CP fiber phases have been employed in highly efficient EV isolations via high‐performance liquid chromatography (HPLC) [ 32 – 34 , 38 , 39 ] and solid‐phase extraction (SPE) tip [ 35 – 37 , 40 ] formats, concentrating on what would be called analytical‐scale processing.…”

“…Marcus and colleagues have developed a hydrophobicity‐based EV isolation method employing capillary‐channeled polymer (C‐CP) fiber stationary phases to address the shortcomings of the currently available EV isolation methods [ 32 , 33 , 34 , 35 , 36 , 37 , 38 , 39 , 40 ]. These C‐CP fiber phases have been employed in highly efficient EV isolations via high‐performance liquid chromatography (HPLC) [ 32 – 34 , 38 , 39 ] and solid‐phase extraction (SPE) tip [ 35 – 37 , 40 ] formats, concentrating on what would be called analytical‐scale processing. In both cases, the isolation of EVs is driven by an organic modifier‐assisted hydrophobic interaction chromatography (HIC) solvent system, where EVs have been obtained from several complex biofluids, including urine, saliva, blood serum, cervical mucus, and CCM from Dictyostelium discoideum cell culture [ 32 , 35 , 39 ].…”

Determination of the Loading Capacity and Recovery of Extracellular Vesicles Derived from Human Embryonic Kidney Cells and Urine Matrices on Capillary-Channeled Polymer (C-CP) Fiber Columns

“…To address the prevalent limitations of available isolation methods, Marcus and coworkers [23][24][25][26][27][28][29][30][31][32] developed a hydrophobic interaction chromatography (HIC) method for the isolation and purification of EVs from a variety of complex biological matrices. The HIC method is performed on polyester (PET) capillary-channeled polymer (C-CP) fiber stationary phases, providing unique benefits in terms of low solvent transport impedance and highly efficient solute mass transfer versus other chromatographic stationary phases [33][34][35][36][37][38].…”