A biosensor was developed using glutathione peroxidase (GSH-Px) as the enzyme. Firstly, platinum (Pt) nanoparticles were deposited onto a glassy carbon paste electrode (GCPE), and then GSH-Px was immobilized by means of gelatin that was then crosslinked with glutaraldehyde. The measurement was based on the electrochemical oxidation of GSH to its disulfide form in the presence of hydrogen peroxide. The linear range was found to be between 10 and 250 M, with a correlation coefficient of R-2 = 0.9968. The R.S.D value for 25 M GSH (n = 6) was calculated as 2.92%. Finally, the proposed biosensor was used to analyze GSH in a synthetically prepared plasma sample, and a promising recovery value was obtained.