Abstract
Extracellular vesicles (EVs) are membranous vesicles secreted by both prokaryotic and eukaryotic cells and play a vital role in intercellular communication. EVs are classified into several subtypes based on their origin, physical characteristics, and biomolecular makeup. Exosomes, a subtype of EVs, are released by the fusion of multivesicular bodies (MVB) with the plasma membrane of the cell. Several methods have been described in literature to isolate exosomes from biofluids including blood, urine, milk, and cell culture media, among others. While differential ultracentrifugation (dUC) has been widely used to isolate exosomes, other techniques including ultrafiltration, precipitating agents such as poly-ethylene glycol (PEG), immunoaffinity capture, microfluidics, and size-exclusion chromatography (SEC) have emerged as credible alternatives with pros and cons associated with each. In this review, we provide a summary of commonly used exosomal isolation techniques with a focus on SEC as an ideal methodology. We evaluate the efficacy of SEC to isolate exosomes from an array of biological fluids, with a particular focus on its application to adipose tissue-derived exosomes. We argue that exosomes isolated via SEC are relatively pure and functional, and that this methodology is reproducible, scalable, inexpensive, and does not require specialized equipment or user expertise. However, it must be noted that while SEC is a good candidate method to isolate exosomes, direct comparative studies are required to support this conclusion.
Highlights
Extracellular vesicles (EVs) can be largely divided into three main subtypes: apoptotic bodies, microvesicles, and exosomes, and are classified based on their cellular origin, physiochemical, and biomolecular properties [1] (Figure 1)
There is a lack of protocol standardization in exosome isolation methods
This void has led to the development of advanced pioneering techniques in order to optimize exosome isolation from a variety of biological fluids [114]
Summary
Extracellular vesicles (EVs) can be largely divided into three main subtypes: apoptotic bodies, microvesicles, and exosomes, and are classified based on their cellular origin, physiochemical, and biomolecular properties [1] (Figure 1). The largest of these EVs, apoptotic bodies arise from the outward blebbing of an apoptotic cell membrane, resulting in phosphatidylserine-rich vesicles 500–5000 nm in diameter. Exosomes are the smallest EVs (30–150 nm) and are formed by the exocytosis of multivesicular bodies (MVBs) liberating intraluminal vesicles upon fusion with the plasma membrane [2] The biogenesis of these intraluminal vesicles occurs through endosomal sorting complexes required for transport (ESCRT) or a soluble N-ethylmaleimide sensitive factor (NSF) attachment protein receptor (SNARE)-based system. Exo-spinTM mini-column, qEV single Sephacryl S-500 HR qEV Exo-spinTM column qEV original qEV70s single, Illustra Sephacryl S-1000 qEV original, Sepharose CL-2B, Sepharose CL-4B
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