Human urinary extracellular vesicles protein LC-MS/MS

Urinary extracellular vesicles (uEV) are a precious new source of biomarkers reflecting the physiological state of the upstream parent cells and organ systems. However, urinary Tamm-Horsfall Protein (THP; uromodulin) interferes heavily with all vesicle isolation attempts, precipitates with normal urinary proteins thus representing an unwanted “contaminant” in urinary assays. Here we developed a simple method to manage THP efficiently. Accordingly, uEV were isolated by hydrostatic filtration dialysis (HFD) and treated with a solution of urea to release most uEV. Presence of uEV was then confirmed by transmission electron microscopy, Western blot and proteomic profiling in MS, all showing that urea treatment did not affect integrity of uEV.