© 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim We demonstrate the guiding principles behind simple two dimensional self-assembly of MOF nanoparticles (NPs) and oleic acid capped iron oxide (Fe3O4) NCs into a uniform two-dimensional bi-layered superstructure. This self-assembly process can be controlled by the energy of ligand–ligand interactions between surface ligands on Fe3O4NCs and Zr6O4(OH)4(fumarate)6MOF NPs. Scanning transmission electron microscopy (TEM)/energy-dispersive X-ray spectroscopy and TEM tomography confirm the hierarchical co-assembly of Fe3O4NCs with MOF NPs as ligand energies are manipulated to promote facile diffusion of the smaller NCs. First-principles calculations and event-driven molecular dynamics simulations indicate that the observed patterns are dictated by combination of ligand–surface and ligand–ligand interactions. This study opens a new avenue for design and self-assembly of MOFs and NCs into high surface area assemblies, mimicking the structure of supported catalyst architectures, and provides a thorough fundamental understanding of the self-assembly process, which could be a guide for designing functional materials with desired structure.