Updated project metadata. In this study, we aimed to investigate if the removal of abundant plasma proteins prior to EV isolation could improve plasma-derived EV characterization by LC-MS/MS by improving the proteome coverage. Plasma depletion was performed using single-use spin columns prior to SEC-based EV isolation. Afterwards, EVs derived from undepleted and depleted plasma were characterized by mass spectrometry (MS)-based proteomics using a data-independent acquisition (DIA) approach.