Extracellular vesicles (EVs) are a diverse group of nano-sized lipid membrane-enclosed particles that are shed by all cells in the body and can provide a unique signature of their cells of origin, including tumor cells. The aim of this study was to characterize circulating EVs in serum from high grade serous ovarian carcinoma (HGSOC) patients compared to healthy controls to identify novel prognostic biomarkers and therapeutic targets. EVs in the serum of HGSOC patients (n=4) and healthy controls (n=5) were isolated using membrane affinity spin columns followed by label-free liquid chromatography-tandem mass spectrometry (LC-MS/MS) to identify differentially expressed proteins. Selected candidate EV proteins that were increased in HGSOC serum (KRT4, LTBP2, MARCKS, SPP1/OPN, and UCHL1) were explored further using online databases and independent patient tissue cohorts. mRNA expression and protein levels of KRT4, MARCKS, SPP1, and UCHL1 were increased in HGSOC compared to normal tissues. Increased mRNA expression of all candidate EV proteins was associated with poor patient outcome. Immunohistochemistry analysis showed higher stroma UCHL1 level was associated with reduced progression-free survival. The UCHL1 inhibitor, LDN-57444, reduced cell metabolic activity of ovarian cancer cell lines (OVCAR3, CaOV3, COV362, OAW28, and A2780) and in primary ovarian cancer cells with high UCHL1 levels. LDN-57444 also blocked motility and invasion of OVCAR3 cells and promoted apoptosis in HGSOC explant tissues with high UCHL1 abundance. The EV biomarker UCHL1, has the potential to be used as a novel prognostic and therapeutic target for HGSOC.