Physical properties, such as density, refractive index and morphology of the aerosol particles will be changed when the relative humidity of the environment is relatively high, which leads to a significant variation of the scattered light signal collected by the single particle light scattering measurement system. If the calibration parameters in the condition of low humidity are applied to inverse aerosol mass concentration, the measurement accuracy will certainly be degraded. Therefore, the calibration parameters of the system must be corrected. In this case, the monotonic relationship between the particle size and the scattered light signal collected by the system should be considered carefully. Combining the Mie scattering theory with the humidity growth model of the particle size and the refractive index, the influence of relative humidity on the scattered light signals of spherical particles has been investigated in this paper. The results indicate that the diameters of spherical particles grow exponentially and the refractive index decreases linearly when the relative humidity is increased between 60% and 95%. Consequently, a significant change of the angular distribution of the scattering intensity occurs. The forward-scattered light is getting stronger, while the back-scattered light becomes weaker. Meanwhile, the scattered light flux of the particle within a solid angle displays exponential growth with increasing the relative humidity. Thus, the monotonous relationship between the particle size and the scattered light flux has changed significantly in the case of the high humidity, and their quadratic coefficient decreases clearly assuming the particle density invariant, which causes an obvious increase of the calibrated proportional coefficient. The results of this work provide an important theoretical guidance for the humidity correction of calibration parameters for the inversion of aerosol mass concentration by the single particle light scattering method.