Abstract: The microscopic imaging technology based on microspheres has been popular in recent years due to its advantages of being simple and free-labeled. However, there is a lack of analytical research on how imaging parameters influence imaging resolution ability. A method for analyzing the imaging resolution ability based on the point source imaging is proposed. The influence of the microsphere size, the distance between the microsphere and the point source on the imaging resolution ability is analyzed by simulations. The results demonstrate that the imaging resolution ability decreases with the increase of the microsphere size. The imaging resolution ability also shows a downward trend with the increase of the distance between the microsphere and the point source when the distance is less than 800 nm. When the distance is more than 800 nm, the imaging resolution ability doesn’t decrease significantly. The simulation results could guide experiments based on microspheres. The proposed method provides the theoretical basis for the research work on the simulation analysis of the imaging resolution ability of microspheres.