Low Energy Electron Holography
and in-situ Electrospray Ion Beam Deposition
 

The lack of a single-molecule imaging method for structurally complex and three-dimensional molecules challenges the community to advance current methods and explore new approaches. In our laboratory, we are developing a low-energy electron holographic microscope (LEEH) capable of imaging of single biomolecules, such as proteins, protein complexes, and nucleic acids, as well as organic molecules and inorganic nanometer-sized objects, with sub-nm spatial resolution and millisecond time resolution over a wide temperature range (30K to 500K).

The powerful combination of the novel in-situ electrospray ion beam deposition machine with the unique characterization method of LEEH enables the imaging of single biomolecules, with 5 Ångstrom resolution. In‑situ electrospray ion beam deposition features quadrupole mass selection, mass spectrometry, ion mobility filtering (conformational selection) and full spatial-, energy- and current characterization of the ion beam. It allows the preparation of contamination-free samples of native biomolecules deposited on a single layer of graphene, which can be imaged using LEEH microscopy. LEEH uses a coherent low energy electron beam (kinetic energies of 30-250eV) to image the individual biomolecules without imposing radiation damage. Thus, in contrast to structure determination by X‑ray diffraction or in cryo‑electron microscopy, no averaging over several different molecules is required, allowing the investigation of flexible domains and flexible regions that remain elusive to ensemble averaging methods.

Operating the microscope at low temperatures and in a low vibration environment is in progress to push the resolution of the microscope to the atomic scale. Moreover, the temporal resolution of the imaging is in the range of 10 - 100 ms and is therefore suitable to access dynamic properties or conformational changes of individual bio/macromolecule, providing insights into biomolecular processes.