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Abstract
Single-particle diffractive imaging is one of the key foundational goals behind the establishment of X-ray Free-Electron Laser (XFEL) facilities. Outrunning radiation damage, extremely intense femtosecond XFEL pulses open up the possibility of imaging uncrystallized aperiodic single-particles frozen in time at room-temperature at the timescales of atomic and electronic motions and thus enabling the capturing of complete energy landscape of molecules both at ground and excited states with sufficiently large data. Despite the current sample-delivery and background scattering challenges, there has been a steady progress in XFEL-single-particle imaging (XFEL-SPI), especially with large symmetric viruses. As a step towards XFEL imaging of single-macromolecules and small-proteins, here we report the coherent diffractive imaging of 3D DNA-origami molecular scaffolds using the soft-X-ray pulses at the European X-ray Free-Electron Laser (EuXFEL). Asymmetric and symmetric 3D DNA-origami scaffold structures were nebulized and delivered to the XFEL beam using an aerodynamic lens stack. The aerosolized DNA-origami structures were intact, and tens-of-thousands of diffraction patterns with expected size and shape matching the simulations of corresponding cryo-EM structures were collected, which were 3D mergeable in the reciprocal space with the expand-maximize-compression (EMC) algorithm. This first demonstration of imaging intact electrospray-aerosolized 3D DNA-origami structures is an important first step towards exploring the application of DNA-origami in XFEL diffractive imaging, especially as, but not-limited-to, molecular scaffolds for small proteins, as references in holographic single-particle imaging or as carriers of strongly-scattering nanoparticle references, and also in time-resolved diffractive imaging of photonic/photoactivatable DNA-origami machines.