was at an applied potential of +0.28 V versus reference Ag/AgCl electrode at ambient temperature. Experimental results show that the current output of the biosensor and the ALT concentration maintained an excellent linear relationship over the ALT concentration range of 0 to 544 ng/mL. Furthermore, the measurement from the biosensor agreed very well with the clinical “gold standard” spectrophotometric results of ALT in human serum. Also, for the analysis of potentially high ALT concentration, a 10-fold dilution of the sample (human serum) can first be carried out. The biosensor can detect the ALT at lower concentration accurately. Thus this single use, disposable and relatively inexpensive ALT biosensor can be very useful for clinical and point-of-care applications.