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Potentiometric biosensors based on flexible arrayed silver paste electrode and copper-doped zinc oxide sensing film modified by iron-platinum nanoparticles (FePt NPs) are designed and manufactured to detect lactate in human. The sensing film is made of copper-doped zinc oxide (CZO) by a radio frequency (RF) sputtering system, and then modified by iron-platinum nanoparticles (FePt NPs). The surface morphology of copper-doped zinc oxide (CZO) is analyzed by scanning electron microscope (SEM). FePt NPs are analyzed by X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FTIR). The average sensitivity, response time, and interference effect of the lactate biosensors are analyzed by voltage-time (V-T) measurement system. The electrochemical impedance is analyzed by electrochemical impedance spectroscopy (EIS). The average sensitivity and linearity over the concentration range 0.2–5 mM are 25.32 mV/mM and 0.977 mV/mM, respectively. The response time of the lactate biosensor is 16 s, with excellent selectivity.