Coherent X-ray diffractive imaging of biological samples in 2D and 3D with synchrotron and XFEL radiation

Coherent X-ray diffractive imaging (CXDI) is a novel microscopy technique that uses the unique X-ray properties of advanced synchrotron and X-ray free-electron laser (XFEL) radiation sources. According to Abbe's theory, the wavelength of light sets the ultimate limit of resolution in microscopes. With X-rays the limit is at the order of a few nanometer or less, well suited for high resolution microscopy.Two major obstacles complicate lens-based conventional X-ray microscopy: Low contrast, as a result of the weak interaction of hard X-rays with matter and the lack of efficient image forming optics. The contrast of unstained specimen can be improved by the choice of the wavelength of only a few nanometer. However, the transparency of matter under X-ray illumination gives insight into the inner structure without slicing the specimen into thin sections. Lenses can be replaced entirely by computational phase retrieval algorithms which are at the heart of CXDI. In CXDI coherent diffraction patterns are measured and converted into images of the object under coherent illumination.Biological objects can be observed with the standard methods of visible light (VIS) microscopy. Structural biology uses sub-nanometer resolution crystallographic methods. CXDI is the technique that can address the resolution range between 200nm to 1nm which is otherwise difficult to reach with VIS microscopy or crystallographic methods. In this thesis methodological improvements in CXDI of biological samples are presented. Single particle imaging in three dimensions (3D) of a virus of 70 nm size using CXDI at LCLS XFEL sources is described, from raw data selection to the final 3D virus density reconstruction. This result represents the to date highest resolution (below 10 nm) virus reconstruction without imposing any symmetry constraints on the measured data. For larger specimen, on the order of 50 μm, the ptychographic CXDI method using the PETRA III synchrotron radiation source is described. Major highlights obtained in the research presented here are the coherence retrieval and quantitative and non-destructive imaging results at 54 nm resolution of a biological cell specimen obtained with ptychography.