Analysis of exosome purification methods using a model liposome system and tunable-resistive pulse sensing

Abstract

Exosomes are vesicles which have garnered interest due to their diagnostic and therapeutic potential. Isolation of pure yields of exosomes from complex biological fluids whilst preserving their physical characteristics is critical for downstream applications. In this study, we use 100 nm-liposomes from 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) and cholesterol as a model system as a model system to assess the effect of exosome isolation protocols on vesicle recovery and size distribution using a single-particle analysis method. We demonstrate that liposome size distribution and ζ-potential are comparable to extracted exosomes, making them an ideal model for comparison studies. Four different purification protocols were evaluated, with liposomes robustly isolated by three of them. Recovered yields varied and liposome size distribution was unaltered during processing, suggesting that these protocols do not induce particle aggregation. This leads us to conclude that the size distribution profile and characteristics of vesicles are stably maintained during processing and purification, suggesting that reports detailing how exosomes derived from tumour cells differ in size to those from normal cells are reporting a real phenomenon. However, we hypothesize that larger particles present in most purified exosome samples represent co-purified contaminating non-exosome debris. These isolation techniques are therefore likely nonspecific and may co-isolate non-exosome material of similar physical properties.

Therapeutic biomaterials based on extracellular vesicles: classification of bio‐engineering and mimetic preparation routes

“…Comparative results of UC are difficult to obtain due to the high number of models available in the market, and this could affect the quality of the isolated product [43]. The use of a fixed instrument would keep low variability between batches [47]. …”

Therapeutic and diagnostic applications of extracellular vesicles

“…Additionally, the impact of the high shear forces on the vesicle integrity are under debate. While some studies indicate no changes in the integrity of the EVs after UC [68,69] others show subtle EV aggregation influencing the EV biodistribution [70,71].…”

Tunable Resistive Pulse Sensing: Potential Applications in Nanomedicine

“…As a result, there have been a surge in the number of diagnostic modalities to study EVs. Over the last few years, a number of studies have employed TRPS as a diagnostic tool to characterise EVs [48][49][50][51][52].…”

“…Lane et al [50] studied the effects of various exosome isolation protocols on vesicle recovery and size distribution using TRPS and 100nm liposomes as a model system, shown in figure 5. They demonstrated that liposome size distribution and ζ-potential are comparable to those of extracted exosomes, making them an ideal model for comparison studies.…”