Rapid and high yield isolation of extracellular vesicles with purity by application of size exclusion fast performance liquid chromatography

Extracellular Vesicles (EVs) have emerged as potential biomarkers for diagnosing a range of diseases without invasive procedures. Extracellular vesicles also offer an advantage compared to synthetic vesicles, for delivery of various drugs. However, limitations in segregating EVs from soluble proteins have led to inconsistent EV retrieval rates with low levels of purity. Researchers at the University of Texas MD Anderson Cancer Center have developed a new high-yield (>95%) and rapid (<20 min) EV isolation method called Size Exclusion Fast Performance Liquid Chromatography (SE FPLC). The researchers show SE FPLC can effectively isolate EVs from multiple sources including EVs derived from human and mouse cells and serum. The results indicate that SE FPLC can successfully remove highly abundant protein contaminants such as albumin and lipoprotein complexes, which can represent a major hurdle in large scale isolation of EVs for clinical translation. Additionally, the high-yield nature of SE FPLC allows for easy industrial upscaling of extracellular vesicles production for various clinical utilities. Moreover, SE FPLC enables analysis of very small volumes of blood for use in point-of-care diagnostics in the clinic. Collectively, SE FPLC offers many advantages over current EV isolation methods and offers rapid clinical utility potential.

The size exclusion – fast protein liquid chromatography EV isolation system: SE-FPLC

(A-E) Schematic overview of the workflow. (A) A heterogeneous concentrated conditioned media sample mixture is loaded onto a FPLC compatible syringe. (B) After loading the concentrated conditioned media onto the SE-FPLC column EVs are separated from small biomolecules using the AKTA Pure chromatography system. (C) A built-in UV monitor is used to perform A280nm measurements. The eluting fractions corresponding to each peak are collected via an automated fraction collector. (D) Fractions of interest are subsequently concentrated using a concentration filter. (E) Purified EVs are stored at -80 oC until further usage and downstream analysis.