ABSTRACT
The study of EVs is currently a hot topic. Their discovery dates back to 19674 in the case of MVs and 19815 for exosomes, and their description as tools of intercellular communication was proposed in 1984.6 Cells release exosomes constitutively and MVs or apoptotic bodies after activation or apoptosis, respectively. When this happens, the vesicles drift around in the extracellular space, where they could be broken down or migrate to distant places by diffusion, appearing in different biological fluids like saliva, urine, or blood. Their functions are yet to be fully established, but key roles could include antigen presentation and immunostimulatory or inhibitory activities by exosomes, procoagulant activity, contribution to tumor invasiveness, the pathogenesis of rheumatoid arthritis, induction of oncogenic cellular transformation, and fetomaternal communication.7 The investigation of EVs is being performed from different perspectives. To understand how and why they perform their functions, one approach is proteomic technology to study the cellular machinery they may contain, that is, proteins. The improvement of mass spectrometry (MS) abilities in recent decades has promoted the use of these techniques in many research fields, and one, where the interest is growing fastest, is the study of MVs. As with other applications of proteomics, the first studies aimed to map the proteome of different subsets of EVs in healthy subjects and/or cell lines (for a recent review see Raimondo et al.8). Currently, studies are diversifying, and the interest is shifting toward the comparison of healthy and disease states with the aim of finding biomarkers of these diseases where MVs play important roles.7 A prerequisite for successful proteomic analysis is a simple and robust sample preparation method.9 The field of blood EV proteomics has been hindered by a lack of standardized protocols to obtain EVs from blood. Previous chapters of this book have dealt with various analytical methods applied to EVs. However, there are two issues related to their isolation that are very relevant for proteomic analyses: the EV preparation should not contain any contaminants (e.g., cells or plasma proteins), and the amount of protein obtained should be sufficient for the analysis. In the next section we will highlight some recommendations for blood-derived EV isolation for proteomic analysis.
