A label-free electrochemical sensor is reported in the present work for the analysis of activase. Firstly, Au elec- trode was modified by electrochemical deposition of gold nanoparticles to increase the surface area of the Au electrode, which resulted in capturing large amounts of other modifiers. Next, the modified electrode with gold nanoparticles was coated with 3-mercaptopropionic acid (MPA) and L-lysine (L-Lys) amino acid via self-as- sembled monolayers (SAMs). The morphologies of the Au nanoparticles and the fabrication process of the sensor were characterized by scanning electron microscopy (SEM), cyclic voltammetry (CV), and electrochemical im- pedance spectroscopy (EIS), respectively. Under the optimal conditions, this sensor displayed an excellent corre- lation between the charge transfer resistance changes and the amount of the absorbed activase protein, and also demonstrated a good linear range from 0.2 to 1.0 ng mL −1 of activase, with a detection limit of 0.01 ng mL −1 . The present sensor has the advantages of simplicity (preparing a self-assembled monolayer), cheapness (not using antibodies), fast detection (with an incubation time of 15 min), and good sensitivity.