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Abstract: The host-guest chemistry of cucurbit[8]uril (CB[8]) and the layer-by-layer self-assembly technique are combined to obtain a molecular imprinted polyelectrolyte multilayer film for the recognition and binding of a guest molecule. Cucurbit[8]uril as a ready-made binding site is first associated with a polyelectrolyte and then assembled into a polyelectrolyte multilayer film via layer-by-layer deposition. A cationic guest is subsequently included into the nanocontainer due to specific host-guest interactions. The quantitative analysis of both CB[8] and the included guest molecule in dependence of the surface charge of the multilayer film identifies a high nanocontainer density as well as good to excellent binding efficiencies, therefore yielding a promising imprinted nanomaterial with potential applications in filtration or sensor technology. The investigation of the guest molecule uptake kinetics reveals two processes on different time scales, respectively, which are again related to the charge of the multilayer film surface. The combination of the results obtained from both ultraviolet spectroscopy and dissipative quartz crystal microbalance enables us to describe a full picture of several simultaneous processes initiated by the guest molecule