Considerable attention has been paid to nanoparticle (NP) research because of their fascinating properties and potential applications in nanotechnology and biotechnology. Asymmetrically functionalizing NP is of particular interest because it could directly lead to controlled patterning of NPs for a variety of applications. In this article, we report using 2-dimensional poly(ethylene oxide) (PEO) lamellar single crystals to create a patterned functional (thiol) surface and to immobilize gold NPs (AuNPs). We demonstrate that patterning AuNPs could be achieved by incubating these single crystals with gold sol and the AuNP areal density could be easily controlled by polymer molecular weight as well as the incubation time. Melting and recrystallization of the AuNP-covered PEO single crystals led to dewetting of PEO. AuNP chains were also observed during the recry stallization process, which was attributed to the dendritic growth of edge-on PEO crystals. Furthermore, this unique technique also enables asymmetric functional i zation of AuNPs. Free-standing bilayer AuNP/PEO films were obtained. Dissolving PEO single crystals led to free asymmetrically functionalized AuNPs and AuNP complexes. This approach provides a novel means to pattern AuNPs and synthesize asymmetrically functionalized AuNPs. We also anticipate that this methodology could be applied to other metal or semiconductor NPs.