Cyclic tangential flow filtration system for isolation of extracellular vesicles

Abstract

Size-based filtration techniques have been developed for high-throughput isolation of extracellular vesicles (EVs). Conventional direct filtration systems have limitations in that large particles generally not only block the pores of the membrane but also damage the particles because of the high fluid pressure. Here, we propose a cyclic tangential flow filtration (TFF) system that includes two membranes with pore sizes of 200 and 30 nm, connected to a peristaltic pump that feeds the stream flowing to the membrane for continuous circulation. The cyclic TFF system is better able to isolate the specific 30–200 nm size range in one step through dual cyclic filtration compared with direct filtration (DF) and single cyclic TFF (scTFF). We further introduced a buffer-exchange process to the dcTFF system after filtration to remove contaminants for more efficient purification. As a result of comparative evaluation of dcTFF and ExoQuick, EVs isolated by dcTFF had more abundant exosome markers and active EVs. The cyclic TFF system not only has great potential to separate EVs with high selectivity and separation efficiency in small volumes of samples but can also be used in clinical applications, including medical diagnostic procedures.

Methodologies to Isolate and Purify Clinical Grade Extracellular Vesicles for Medical Applications

“…These kits are easy to use, have a high yield rate and can be used for different types of bio-fluid samples [ 64 , 65 ]. Other methods are tangential flow filtration (TFF), direct filtration, and cyclic TFF [ 66 ].…”

Application of engineered extracellular vesicles for targeted tumor therapy

“…Furthermore, the employment of ultrafiltration membranes and columns with pore diameters of varying size allows for creating a population of particular dimensions, and plasma EVs of specific diameters have been obtained using such columns [ 47 , 48 ]. Notably, however, ultrafiltration is a time-consuming process, the pores in the ultrafiltration membrane are easily clogged, and the pressure applied during isolation may deform or rupture the EVs [ 49 ]. Sequential centrifugation has been proposed as a means of improving the ultrafiltration method.…”

Extracellular Vesicles as a Therapeutic Tool for Kidney Disease: Current Advances and Perspectives

“…The different sizes of EVs have been used for designing novel isolation protocols. Examples such as the ultrafiltration system and tangential flow filtration (TFF) use membranes with a defined molecular weight cut-off ranging from 10 to 100 kDa and have been used to isolate EVs from urine-derived cell culture medium [125,126]. Another method is the size-exclusion chromatography (SEC) that separates EVs from different body fluids based on their hydrodynamic volume and molecular size [128].…”