Many cell types release exosomes and microvesicles (EMVs) with a size range from 0.05-5 μm, harbouring receptors, bioactive and signaling proteins, molecular mediators and nucleic acids for cell-to-cell communication. Microvesicles bud directly from the plasma membrane, in contrast to exosomes which are derived from multivesicular bodies within the cell. Increased levels of EMVs have been observed in plasma, urine and other body fluids in patients suffering from a wide range of common complex diseases, including vascular, metabolic, lung, autoimmune and neurodegenerative diseases, chronic inflammation and cancer. EMVs may affect target cells directly by surface-bound ligands, transferred surface receptors and membrane-associated enzymes, such as glycosylphosphatidylinositolanchored proteins, or delivered cytoplasmic or membrane-associated constituents, such as cytosolic proteins, micro/mRNAs, bioactive lipids and even mitochondria. The use of EMVs as diagnostic markers for the prediction, diagnosis, therapy monitoring and prognosis of complex diseases is becoming increasingly attractive. Novel technologies for analysis of the size, density and molecular composition of EMVs are currently emerging together with methods for their improved isolation and purification out of heterogenous vesicle populations. In addition, the recent revolution in mass-spectroscopy, (micro-) flow cytometry, atomic force microscopy, nanoparticle tracking and biosensing will considerably facilitate the quantitative and qualitative analysis of all the constituents assembled in EMVs. Technologies will be preferred that provide signatures specific for EMV subsets rather than a single or a few parameter(s) averaged for the total EMV population. Those EMV signatures have to be correlated to specific disease states along cross-sectional and longitudinal clinical studies. Moreover, it has to be tested which signatures and molecular components, i.e. EMV subspecies, are most informative to obtain actionable disease information. Ultimately, the reliable, rapid and low-cost analysis of EMVs will support systems biology-based approaches for the diagnosis and therapy of complex diseases and supplements the analytical power of conventional biomarkers.