Cryo soft X-ray tomography (Cryo-SXT) is a valuable tool for high-resolution three dimensional (3D) imaging of cells in near-native preservation state. Cryo-immobilization of cells based on rapid cryogenic freezing is crucial to obtain good imaging in Cryo-SXT. In order to acquire high contrast image of live cells, it need to prepare the specimen ice as thin as possible. However, as the thickness of the ice layer is further reduced, the live cells may be destroyed during the tilt series data collection of 3D imaging due to radiation. Thus, it is essential to seek out a proper thickness range of the ice layer to get high contrast image without radiation damage of cells. For further understanding the relationship between thickness of ice layer and degree of radiation damage, yeast cells with different thickness of ice layer were imaged using Cryo-SXT. Finally the ice layer with thickness about 10 μm was confirmed to be proper for good imaging.