Membrane microvesicles mediate the transfer of biologically active molecules, thus play a crucial role in intercellular communication as well as in the regulation of immune and regenerative processes. Their molecular composition reflects both the cellular origin and the physiological or pathological state of the parent cells, making them potential biomarkers for the early detection of disease, monitoring cancer progression, and evaluating treatment succes. Furthermore, their molecular composition can provide insight into the target cells to which they are directed. In various diseases, both the concentration and molecular composition of membrane microvesicles, including specific protein and other molecular markers, are altered, enabling detection between different pathological conditions and prediction of disease outcomes. Despite their considerable diagnostic and therapeutic potential, the isolation of membrane microvesicles remains a major challenge due to vesicle heterogeneity, the presence of non-target particles, and the lack of standardized isolation protocols. The most commonly used isolation methods include ultracentrifugation, ultrafiltration, and size-exclusion chromatography. In recent years, additional techniques have been developed to improve the specificity of isolated products. Among the most promising techniques are tangential flow filtration combined with size-exclusion chromatography, ultracentrifugation with flow cytometry, batch ultrafiltration, and microfluidic technologies which still requires further optimization to enable large-scale application and efficient separation of extracellular vesicle subpopulations. In most studies, combining multiple isolation methods has resulted in improved purity, higher yield, sepparation of subpopulations and quality of isolated microvesicles. Further optimisation of characterization methods is also required to improve reliability of isolation results. Continued research will be essential for the standardization of isolation protocols and the broader implementation of membrane microvesicles in diagnostic and therapeutic applications.
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