Extracellular Vesicles (EVs) play important roles in a broad range of physiological and pathological processes. The size, number, membrane composition and contents of EVs are highly heterogeneous, dynamic and dependant on the cellular source, state and environmental conditions. Thus, understanding the biophysical diversity in EV population is paramount for linking the impact of EV properties to its biological role and function.
Izon has developed an integrated methodology that utilizes simple Size Exclusion chromatography (SEC) based qEV columns for rapid EV purification from a variety of biological fluids (urine, serum, plasma, CSF, cell culture etc.) followed by accurate, high-resolution particle-by-particle EV biophysical characterization (size, size range, concentration and surface charge) through the qNano that utilizes Tunable Resistive Pulse Sensing (TRPS) technology. The SEC columns provide a convenient, reproducible and highly effective means of eliminating >99% of non-vesicular protein from biological fluid samples. The SEC derived vesicles are isolated within 15 minutes, the process is gentle and does not require addition of agents that induce precipitation or aggregation.
SEC derived vesicles are typically amenable for downstream biological analysis without further purification or concentration. Magnetic particles conjugated with antibodies to CD9 and CD63 reacted with the SEC derived EVS whereas control IgG reactions were negative. The resulting EV-Magnetic complex (immunoprecipitate) were analyzed by TRPS and the size of the complexes verified a proportional increase in size and volume over a defined dose-range. Changes in physical properties indicated the positive selection of specific sub-population.
EV analysis is challenging for conventional technologies because of the small particle sizes of many of these structures. TRPS brings to the EV field a powerful tool for particle concentration and size analysis down to 40nm.