Abstract: Fiber optic oxygen sensor based on fluorescence quenching is developed and its test system is assembled. High-brightness blue LED is adopted as light source. Photomultiplier tube is used as detector. Grating spectrometer is used as spectral instrument and the SR830 lock-in amplifier is used to detect the phase shift between the fluorescence and excited light in this system. The dependence of phase shift on modulated signal frequency and its duty cycle are studied. The experimental results show that the rectangular modulated signal whose frequency is 60 kHz and duty cycle is 10% enabled the sensor detecting precision to achieve maximum. It is very significant for the further study of the optical dissolved oxygen sensor.