The chiral structures have displayed some inevitable and fascinating properties in many research fields, such as chemistry, biology, mathematics, and physics. In this Article, we report the use of stepwise glancing angle deposition technology to produce the 3D chiral nanostructures. Through the optimization of deposition parameters (such as the orientation angle of poly styrene spheres (PSs) array, the deposition angle, thickness, and number), a great number of chiral structures have been achieved, and their size depends on the diameter of PS spheres. These chiral structures all can be simulated and predesigned through the use of a 3D geometrical model, which greatly improves the efficiency of this method. In addition, the circular dichroism spectrum shows that these chiral structures own an obvious Cotton effect, indicating their potential application as 3D chiral metamaterials.