Due to the high demand for structural reliability, Structural Health Monitoring (SHM) has drawn intensive attention. Fiber Bragg Grating (FBG) sensor with many advantages has become one of the most promising sensing elements in this area. However, the axial sensitivity, as the main parameter of the FBG sensor, has received little attention. This paper tried to explore the influence of the ply stacking of composite on the axial sensitivity of the FBG sensor. The calibration test, the repeatability test, and the influence test had been carried out in this paper. Thereinto, the theoretical axial strain sensitivity of the FBG sensor (1.06312 pm/με) was obtained by the calibration test. The repeatability test verified that the effect of preparation and operating conditions on the test results could be ignored, while the influence test was used to demonstrate that the three kinds of stacking could affect the axial sensitivities of the FBG sensors. The experimental results showed that: the axial sensitivities of two FBG sensors embedded between 45°/45° layers were equal. But, a difference was observed between the FBG sensors embedded between the layers of 0°/0°, 45°/45°, and 90°/90°. The maximum relative difference was about 2.587%. It indicated that the laying sequence of the FBG sensor influenced the axial sensitivity of the FBG sensor. Subsequently, the ANSYS software was adopted to simulate the deformation distribution of the sensor under a tensile load of 800N. A good agreement was observed between the numerical prediction and experimental results.