Extracellular micro- and nanoscale membrane vesicles produced by different cells progressively attract the attention of the scientific community. They function as mediators of intercellular communication and transport genetic material and signaling molecules between the cells. In the context of keeping homeostasis, the extracellular vesicles contribute to the regulation of various systemic and local processes. Vesicles released by the tumor and activated stromal cells exhibit multiple functions including support of tumor growth, preparation of the pre-metastatic niches, and immune suppression. Considerable progress has been made regarding the criteria of classification of the vesicles according to their origin, content, and function: Exosomes, microvesicles, also referred to as microparticles or ectosomes, and large oncosomes were defined as actively released vesicles. Additionally, apoptotic bodies represented by a highly heterogeneous population of particles produced during apoptosis, the programmed cell death, should be considered. Because the majority of isolation techniques do not allow the separation of different types of vesicles, a joined term “extracellular vesicles” (EVs) was recommended by the ISEV community for the definition of vesicles isolated from either the cell culture supernatants or the body fluids. Because EV content reflects the content of the cell of origin, multiple studies on EVs from body fluids in the context of cancer diagnosis, prediction, and prognosis were performed, actively supporting their high potential as a biomarker source. Here, we review the leading achievements in EV analysis from body fluids, defined as EV-based liquid biopsy, and provide an overview of the main EV constituents: EV surface proteins, intravesicular soluble proteins, EV RNA including mRNA and miRNA, and EV DNA as potential biomarkers. Furthermore, we discuss recent developments in technology for quantitative EV analysis in the clinical setting and future perspectives toward miniaturized high-precision liquid biopsy approaches.