Nanoscale extracellular vesicles (EVs) including exosomes (50-150 nm membrane particles) have emerged as promising cancer biomarkers due to the carried genetic information about the parental cells. However the sensitive detection of these vesicles remains a challenge. Here we present a label-free electrochemical sensor to measure the EVs secretion levels of hypoxic and normoxic MCF-7 cells. The sensor design includes two consecutive steps; i) Au electrode surface functionalization for anti-CD81 Antibody and ii) EVs capture. The label-free detection of EVs was done via Differential Pulse Voltammetry (DPV) and Electrochemical Impedance Spectroscopy (EIS). The working linear range for the sensor was 102-109 EVs/ml with an LOD 77 EVs/mL and 379 EVs/ml for EIS and DPV based detection. A blood-abundant protein, RhD was used for the selectivity test. In order to assess the performance of the biosensor, the level of EVs secretion by the human breast cancer MCF-7 cell line was compared with enzyme-linked immunosorbent assays (ELISA) and Nanoparticle Tracking Analysis (NTA). Designed label-free electrochemical sensors utilized for quantification of EVs secretion enhancement due to CoCl2-induced hypoxia and 1.23 fold increase with respect to normoxic conditions was found.