Alpha-1 antitrypsin (A1AT) is one of the acute phase proteins which are synthesized in the liver. A1AT inhibits the activity of many proteases, but its main task is to protect the lungs from the attack of neutrophil elastase. In an autosomal hereditary disease known as alpha-1 antitrypsin deficiency, the A1AT level in blood serum decreases, increasing the risk of developing emphysema, liver apoptosis, and liver cancer. Thus, the detection of A1AT concentration in blood serum is very important. In this study, an impedimetric biosensor was developed, forming an SAM (self-assembled monolayer) with 4-mercaptophenylacetic acid (4MPA) on the surface of the gold electrode. An A1AT biosensor was constructed using immobilization of an A1AT-specific antibody (anti-A1AT) after activating the carboxyl groups of 4MPA with EDC/NHS. Each immobilization stage was characterized by using electrochemical impedance spectroscopy, cyclic voltammetry, and scanning electron microscopy with energy dispersive X-ray spectroscopy. With the designed biosensor, precise and highly reproducible results were obtained for A1AT concentrations in the range of 100-600 mu g/mL. A1AT detection was also successfully carried out in artificial serum solutions spiked with A1AT.