To study the effect of the luminance gain of a third-generation low-light-level (LLL) image intensifier on the image quality, we compare and analyze the image quality of the output image under different gain conditions to improve the image quality of a third-generation image intensifier. First, based on the theory of the third-generation LLL image intensifier, it is proven that the luminance gain directly affects the image quality of the image intensifier. Then, via signal-to-noise ratio (SNR) and resolution, two important parameters of the image quality evaluation, image quality evaluation system, and experimental equipment, are set up. Finally, the experimental results show that under moonless illumination, when the luminance gain is at the optimal value, the resolution of the output image is increased from 32 lp/mm to 40.3 lp/mm while the field of view is bright and clear. It has been proven that this research has guiding significance in how to obtain the best image quality of LLL night vision instruments by reasonably setting the luminance gain value in the night environment.