A survey to evaluate parameters governing the selection and application of extracellular vesicle isolation methods

Abstract

Extracellular vesicles (EVs) continue to gain interest across the scientific community for diagnostic and therapeutic applications. As EV applications diversify, it is essential that researchers are aware of challenges, in particular the compatibility of EV isolation methods with downstream applications and their clinical translation. We report outcomes of the first cross-comparison study looking to determine parameters (EV source, starting volume, operator experience, application and implementation parameters such as cost and scalability) governing the selection of popular EV isolation methods across disciplines. Our findings highlighted an increased clinical focus, with 36% of respondents applying EVs in therapeutics and diagnostics. Data indicated preferential selection of ultracentrifugation for therapeutic applications, precipitation reagents in clinical settings and size exclusion chromatography for diagnostic applications utilising biofluids. Method selection was influenced by operator experience, with increased method diversity when EV research was not the respondents primary focus. Application and implementation criteria were indicated to be major influencers in method selection, with UC and SEC chosen for their abilities to process large and small volumes, respectively. Overall, we identified parameters influencing method selection across the breadth of EV science, providing a valuable overview of practical considerations for the effective translation of research outcomes.

Proteomic characterization of extracellular vesicles in programmed cell death

“…Other feasible methods include precipitation, membrane affinity, size‐exclusion chromatography (SEC), iodixanol gradient, phosphatidylserine (PS) affinity, and so on [78]. Different methods affect the purity and yield of EVs, which is important to develop an optimal separation scheme for EV‐related research based on surface markers, size, and components [79]. In practice, sequential centrifugation followed by sequential filtering is perhaps the most widely used isolation method for downstream proteomic analysis.…”