Microfocus X-ray source is the core component of micro-computed tomography (micro-CT). This study investigates the relationship between the changes in the focal spot size and intensity of the transmission microfocus X-ray source caused by the lateral diffusion of the electron beam in the target. Results show that, if the density distribution of the electron beam follows a Gaussian distribution, the distribution of the X-ray intensity should also be Gaussian. The standard deviation of the X-ray intensity distribution exactly represents the size of the X-ray focal spot. Furthermore, results show that when the energy deposition of the electron beam in the target reaches 60%, the intensity of the X-ray produced by the target reaches the maximum value, correspondingly. With an increase in the target thickness, the focal spot size of the X-ray source gradually increases; conversely, an increase in the acceleration voltage of the electron beam can appropriately reduce the focal spot size of the X-ray source. This study provides theoretical guidance for target material selection and design of transmission microfocus X-ray source.