细胞外小泡
分离(微生物学)
细胞外
大小排阻色谱法
小泡
化学
色谱法
生物
生物化学
细胞生物学
生物信息学
膜
酶
作者
Ana Gámez‐Valero,Marta Tortajada,Laura Carreras‐Planella,M. Franquesa,Katrin Beyer,Francesc E. Borràs
摘要
Abstract Extracellular vesicles (EVs) have become an attractive field among the scientific community. Yet, a major challenge is to define a consensus method for EVs isolation. Ultracentrifugation has been the most widely used methodology but rapid methods, including Size Exclusion Chromatography (SEC) and/or precipitating agents such as Polyethylene glycol (PEG) or PRotein Organic Solvent PRecipitation (PROSPR) have emerged. To evaluate the impact of these different methods on the resulting EV preparations, plasma EVs were isolated using SEC, PEG and PROSPR and their total protein content, NTA and Cryo-electron microscopy profiles and EV-markers were compared. Also, their effect on recipient cells was tested. Low protein content and Cryo-EM analysis showed that SEC removed most of the overabundant soluble plasma proteins, which were not removed using PEG and partially by PROSPR. Moreover, only SEC allowed the detection of the EV-markers CD9, CD63 and CD81, LGALS3BP and CD5L, suggesting a putative interference of the precipitating agents in the structure/composition of the EVs. Furthermore, PEG and PROSPR-based EV isolation resulted in reduced cell viability in vitro . These results stress that appropriate EV-isolation method should be considered depending on the forthcoming application of the purified EVs.
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