An electrochemical method to determine alanine aminotransferase (ALT) activity over its normal and elevated physiological range was developed based upon detection of l-glutamate at a glutamate oxidase-modified platinum electrode. Measurements were carried out in the presence of ALT co-substrates l-alanine and α-ketoglutarate and current response from either the oxidation of hydrogen peroxide or the re-oxidation of the mediator ferrocene carboxylic acid was employed. The enzyme electrode was tested over a 6-month period and found to retain 79% of its original activity towards ALT detection with >200 measurements performed over this time. Signals associated with interfering electroactive species (ascorbic acid and uric acid) were eliminated using background subtraction at a denatured glutamate oxidase enzyme electrode. The sensitivity of the device was found to be 0.845 nA U −1 L ALT with t 90 = 180 s, linear range 10–1000 U L −1 and LOD of 3.29 U L −1 using amperometry at E app = 0.4 V vs. Ag/AgCl at 308 K (35 °C).