Isolation of extracellular vesicles: Determining the correct approach (Review)

Abstract

The discovery of extracellular vesicles (EVs) has revised the interpretation of intercellular communication. It is now well established that EVs play a significant role in coagulation, inflammation, cancer and stem cell renewal and expansion. Their release presents an intriguing, transporting/trafficking network of biologically active molecules, which are able to reach and modulate the function/behavior of the target cells in a variety of ways. Moreover, the presence of EVs in various body fluids points to their potential for use as biomarkers and prognostic indicators in the surveillance/monitoring of a variety of diseases. Although vast knowledge on the subject of EVs has accumulated over the years, there are still fundamental issues associated with the correct approach for their isolation. This review comprises the knowledge on EV isolation techniques that are currently available. The aim of this review was to make both experienced researchers and newcomers to the field aware that different types of EVs require unique isolation approaches. The realization of this ‘uniqueness’ is the first step in the right direction for the complete assessment of EVs.

Extracellular vesicles from bone marrow‐derived mesenchymal stromal cells support ex vivo survival of human antibody secreting cells

“…The supernatant was aspirated using a sterilized glass Pasteur pipette, with the resulting pellet, which we refer to as EV fractions (Supplemental Figure 1), resuspended in 500–600 µL sterile PBS and stored at −80°C until use. Our method, which did not use a sucrose gradient, was adapted from other similar publications [30,54]. …”

Ceramide and Exosomes: A Novel Target in Cancer Biology and Therapy

“…Immunoprecipitation using antibodies against exosome surface proteins can alleviate this problem and even separate exosomes derived from different cell types. However, the yield is usually lower than with centrifugation techniques and often compromises a quantitative analysis of exosome function (Brett et al, 2017; Momen-Heravi et al, 2013; Nakai et al, 2016; Szatanek, Baran, Siedlar, & Baj-Krzyworzeka, 2015). Recently, great strides were made in the field of exosome isolation introducing new techniques such as ultrafiltration and microfluidics-based separation.…”

Glycosylation of extracellular vesicles: current knowledge, tools and clinical perspectives

“…This research links to a broader question on the impact of isolation methods on EV samples. To elaborate, many comparison studies have been performed to systematically examine the impact of methods such as ultracentrifugation, chromatography, ultrafiltration and commercial isolation kits on the protein and RNA cargoes of EVs [109–111] but these have not addressed exosome glycosylation. Lectin-based purifications could confer a bias in this regard but for other methods the effect of glycosylation on exosome isolation is less certain.…”