Current approaches for exosome processing and the emerging importance of subtyping

Studying exosomes is extremely challenging, due to enormous technical difficulties in defining and isolating such small and molecularly diverse nanovesicles. The progress in the biomedical field has been rather slow because of the lack of effective isolation approach. Extracellular vesicles (EVs) shed by cells are a highly complex population with a size range of 10-1000 nm, which is mainly composed of exosomes and microvesicles that differ in cell origin and biogenesis. Due to the substantial size overlap among these membrane vesicles, confusion on the origin and nomenclature of EVs has spread through the field. This talk reviews both historic and cutting edge exosome isolation technologies and platforms along with exosome methods of production, isolation and purification which have been shown to have significant effects on yield, viability and function. Further, exosome subtypes are critical to future research, biomarker identification and therapeutic development. Currently, the most-recognized isolation approach is ultracentrifugation, which has been shown to exhibit marked heterogeneity in isolated exosomes with variable sizes and protein compositions, indicating the inability to differentiate exosome subpopulations and other vesicle types (e.g., microvesicles and apoptotic blebs). Moreover, the isolation efficiency of ultracentrifugation is rather low (~5-25%) and involves multiple time-consuming steps for repetitive centrifugation and filtration (>10 hrs). Size-exclusion methods are prone to dilution, contaminations, and pressure-caused damage. To date, there are no well-defined methods for exosome isolation in high-efficiency and high-throughput, which severely hindered the broad applications of exosomes in the biomedical field. We close by introducing Clara ExoRelease Beads, a new purification method called “immuno-isolation and release” that offers standardization, reproducibility, and subtyping while preserving exosome functionality.

Learning Objectives:

1. Understand methods of isolating and enriching exosomes along with their pros/cons of the different techniques

2. What do we mean when we talk about subtyping, and what is its importance?