This study builds an intra-pixel response measurement system to overcome the issue arising when the output energy of the point target fluctuates with position, which is caused by the fill factor of the forward-illuminated CMOS image sensor less than 1. According to the interior design structure of the pixel, an intra-pixel response model is established. Two holes with different radii are used to measure the intra-pixel response functions of four pixels in the sensor. The response value of the pixel is recorded when the hole is moved to different positions. The full widths at half maximum of Gauss function of the point target and the intra-pixel response function are calculated. Experimental results show that the spatial distribution characteristics of sensitivity within the pixel can be described by calculating the intra-pixel response function. The measurement and resolution method proposed herein is able to provide a useful reference for the microscopic representation of this type of sensor.