Collision electron spectroscopy (CES) which is based on probe diagnosis and allows for the identification of the impurity gas in pure inert gas has provided a new method for gas analysis. Due to the higher electron temperature in the positive column, the negative glow region is frequently selected as the optimization for gas analysis in DC discharge operated at a relatively high gas pressure. In this paper, the effect of ambipolar diffusion of fast electrons, and the importance of ambipolar diffusion considered as a deep influence on the resolution of fast penning electrons in the positive column, have been verified by designed experiment. It was found that the increase in plasma density has an energy loss effect on fast electrons, resulting a smooth effect on fast electron detection at pressures of about few hundred Pascals.