The rotational performance of the spindle-bearing system has critical influence upon the geometric shape and surface roughness of the machined parts. The effects of preload and preload method on the rotational performance of the spindle-bearing system is explored experimentally to reveal the role of preload and preload method in spindle rotational performances under different speeds. A test rig on which both the rigid preload and elastic preload can be realized, equipped with variable preload spindle-bearing system, is developed. Based on the mechanical model, the relationship of the axial preload and negative axial clearance of the spindle-bearing system is provided. Rotating sensitive radial error motion tests are conducted for evaluating synchronous and asynchronous radial errors of the variable preload spindle-bearing system under different rotating speeds and preload methods. The change regularity of synchronous and asynchronous radial errors with preloads under different rotating speeds are given. The results show that the preload plays an important role on the rotational performance of spindle-bearing system. The rigid preload is more efficient in achieving better rotational performance than elastic preload under the same rotating speed. Furthermore, this article significantly guides the preload designing and assembling of the new spindle-bearing system.