In recent decades, nanospheres have demonstrated unique self-assembly behaviors due to their spherical shape, small size, high specific surface area, and tunable surface properties. These characteristics enable the creation of hierarchical or periodic functional surfaces, which play a critical and distinct role in applications such as optics, wettability regulation, catalysis, and environmental sensing. This chapter provides an overview of the principles, methods, and applications of nanospheres dynamic self-assembly in the development of advanced functional surfaces. First, the interactions in the dynamic self-assembly of nanospheres are discussed, including van der Waals forces, electrostatic interaction, hydrogen bonding, coordination, etc. Next, we introduce the methods for inducing the dynamic self-assembly of nanospheres through solvent evaporation, temperature induction, and external force fields and discuss their advantages and limitations. Then, we enumerate the applications of dynamic self-assembly of nanospheres in developing functional surfaces, including photonic crystals, antireflective coatings, wettability control, environmental monitoring, and so on. Finally, the challenges and future development trends in advancing this technology in the field of nanoscience and technology are discussed, aiming to inspire researchers to continue exploring more exciting possibilities in this field.