We propose a self-assembled DNA origami honeycomb 2D-lattice as a molecular imaging scaffold for Cryo-Electron Microscopy (Cryo-EM). The thin, micrometer-scale supporting scaffold is able to cover extended areas of the holey carbon film and sufficiently resilient to withstand blotting and plunge-freezing forces. Furthermore, the DNA binding sites can be chemically engineered for selective surface-affinity trapping. We demonstrate the advantages of the method to facilitate membrane vesicles sample preparation. This includes increasing the local density of the vesicles, enabling the study of low-abundance membrane complexes, and purifying heterogenous samples by isolating vesicles from cell conditioned medium.