Extracellular vesicles (EVs) are attracting increasing interest with their intrigue role in intercellular communications. Protein phosphorylation in EVs is of great importance for understanding intercellular signaling processes. However, the study of EV phosphoproteomics is impeded by their relatively low amount in limited clinical samples and it is necessary to have a sensitive and efficient enrichment method for EV phosphopeptides. Herein, a novel Ti4+-functionalized and glass-fiber supported hybrid monolithic spin tip, we named PhosTip, was prepared for enriching phosphopeptides from urinary EVs.