**2. The origin and characteristics of extracellular vesicles (EVs)**

Extracellular vesicles (EVs) are produced and released into the extracellular space by all cells. EVs are classified based on differences in their biogenesis, size and functions [2]. The current EV nomenclature recognizes exosomes (30–150 nm), microvesicles (MVs; 150–1000 nm) and apoptotic bodies (>1000 nm). However, within these EV categories, there is considerable heterogeneity. Thus, although all exosomes, now referred to as small EVs (sEVs), originate in multivesicular bodies (MVBs) and thus share the endocytic origin [3], they are divided into tiny exomeres (<35 nm), small exosomes (Exo-S, 50–100 nm) and large exosomes (Exo-L, 100–150 nm) [2]. In our studies of EVs, we consider sEVs derived from MVBs and sized from 50 to 150 nm as exosomes [4] and intermittently refer to them as either exosomes or sEVs. Unlike exosomes, MVs bud off from the surface of parent cells, differ broadly in size and molecular content from exosomes and are called ectosomes or, if they carry oncogenes, oncosomes [2].

#### **Figure 1.**

*Schematic view of an immune capture of tumor-derived exosomes (TEX) using an antibody (capture Ab) specific for an antigen epitope present on the surface of TEX. The capture Ab is labeled with biotin and is co-incubated with exosomes isolated by size exclusion chromatography (SEC) from plasma. These exosomes contain TEX and Non-TEX, which are produced by various non-malignant cells. TEX carry the target Ag and are captured by the Ab. Non-TEX do not carry the target Ag and are not recognized by the capture Ab. Streptavidin-labeled magnetic beads are added to the mix of exosomes and biotinylated Ab. TEX are captured on streptavidin beads and recovered using a magnet. Non-TEX are not captured and remain in solution. Captured TEX are then evaluated for their molecular content by on-bead flow cytometry.*

*Diversity of Extracellular Vesicles (EV) in Plasma of Cancer Patients DOI: http://dx.doi.org/10.5772/intechopen.101760*

During exosome biogenesis, when MVBs filled with intraluminal vesicle fuse with the cellular plasma membrane, exosomes are released into the extracellular space. Due to their endosomal origin, exosomes carry endocytic markers, such as TSG101, ALIX, syntenin-1, flotillin and others but do not contain cytoplasmic proteins, such as calnexin or GRPp94. Importantly, the topography of exosome molecular surface as well as molecular and genetic contents of exosomes resemble those of their parent cells [5]. This similarity of molecular and genetic signatures of tumor-derived exosomes (TEX) to parent tumor cells is the main reason for considering TEX as a "liquid tumor biopsy" [6]. Tumor cells produce large numbers of exosomes, ranging, from 1010 to 1012/mL plasma, and plasma of cancer patients is thus variably but significantly enriched in TEX [7]. However, not only tumor cells but also non-malignant cells in the TME, e.g., immune cells, endothelial cells or cancer-associated fibroblasts (CAFs), produce exosomes, which account for a considerable fraction of total EVs present in cancer patients' plasma [8, 9]. Thus, cancer plasma contains a diverse mix of small and large EVs of various cellular origins and presumably with different molecular content in the vesicle lumen and on the vesicle surface membrane. TEX represent a fraction of all circulating EVs that differs broadly among cancer patients; in patients with malignant melanoma, TEX constitute 20–80% of total plasma EVs (our data), and the ratio of TEX/non-TEX increases with melanoma progression **Figure 1** [7].
