Abstract: The present disclosure provides a three-dimensionally arranged nanoparticle film and a preparation method and use thereof, and belongs to the technical field of nanophotonics. The preparation method includes the following steps: coating nanoparticles with hydrophobic molecules, and dispersing in an organic solvent to obtain a modified nanoparticle dispersion; conducting a surface treatment on a three-dimensional template to obtain a hydrophilic three-dimensional template; and adding water to a surface of the hydrophilic three-dimensional template to form a water film, adding the modified nanoparticle dispersion dropwise on a surface of the water film to conduct self-assembly, and removing the water film to obtain a three-dimensionally arranged nanoparticle film with an array structure.