Three-Dimensional Correlative Imaging of a Malaria-Infected Cell with a Hard X-ray Nanoprobe.

Benefiting from the recent advances of synchrotron X-ray nanoprobes, we demonstrate three-dimensional (3D) correlative nanoimaging on malaria-infected human red blood cells. By combining X-ray fluorescence tomography and phase contrast nanotomography on the same cell with sub-100 nm pixel size, we establish a routine workflow from the data acquisition, data processing, to tomographic reconstruction. We quantitatively compare the elemental volumes obtained with different reconstruction methods, with the total variation minimization giving the most satisfactory results. We reveal elemental correlations in different cell compartments more reliably on reconstructions as opposed to 2D projections. Finally, we determine for the first time the 3D mass fraction maps of multiple elements at the subcellular level. The estimated total number of Fe atoms and the total mass of red blood cells show very good agreement with previously reported values.

Registration of phase-contrast images in propagation-based X-ray phase tomography.

X-ray phase tomography aims at reconstructing the 3D electron density distribution of an object. It offers enhanced sensitivity compared to attenuation-based X-ray absorption tomography. In propagation-based methods, phase contrast is achieved by letting the beam propagate after interaction with the object. The phase shift is then retrieved at each projection angle, and subsequently used in tomographic reconstruction to obtain the refractive index decrement distribution, which is proportional to the electron density. Accurate phase retrieval is achieved by combining images at different propagation distances. For reconstructions of good quality, the phase-contrast images recorded at different distances need to be accurately aligned. In this work, we characterise the artefacts related to misalignment of the phase-contrast images, and investigate the use of different registration algorithms for aligning in-line phase-contrast images. The characterisation of artefacts is done by a simulation study and comparison with experimental data. Loss in resolution due to vibrations is found to be comparable to attenuation-based computed tomography. Further, it is shown that registration of phase-contrast images is nontrivial due to the difference in contrast between the different images, and the often periodical artefacts present in the phase-contrast images if multilayer X-ray optics are used. To address this, we compared two registration algorithms for aligning phase-contrast images acquired by magnified X-ray nanotomography: one based on cross-correlation and one based on mutual information. We found that the mutual information-based registration algorithm was more robust than a correlation-based method.

Holographic imaging with a hard x-ray nanoprobe: ptychographic versus conventional phase retrieval.

We have performed near-field x-ray imaging with simultaneous object and probe reconstruction. By an advanced ptychographic algorithm based on longitudinal and lateral translations, full-field images of nanoscale objects are reconstructed with quantitative contrast values, along with the extended wavefronts used to illuminate the objects. The imaging scheme makes idealizing assumptions on the probe obsolete, and efficiently disentangles phase shifts related to the object from the imperfections in the illumination. We validate this approach by comparison to the conventional reconstruction scheme without simultaneous probe retrieval, based on the contrast transfer function algorithm. To this end, a set of semiconductor nanowires with controlled chemical composition (InP core, insulating SiO2 layer, and indium tin oxide cover) is imaged using the quasi-point source illumination realized by the hard x-ray nanofocus (26 nm×39 nm spot size) of the ID16A Nano-Imaging beamline at the European Synchrotron Radiation Facility.

X-Ray Phase Contrast Tomography Reveals Early Vascular Alterations and Neuronal Loss in a Multiple Sclerosis Model.

The degenerative effects of multiple sclerosis at the level of the vascular and neuronal networks in the central nervous system are currently the object of intensive investigation. Preclinical studies have demonstrated the efficacy of mesenchymal stem cell (MSC) therapy in experimental autoimmune encephalomyelitis (EAE), the animal model for multiple sclerosis, but the neuropathology of specific lesions in EAE and the effects of MSC treatment are under debate. Because conventional imaging techniques entail protocols that alter the tissues, limiting the reliability of the results, we have used non-invasive X-ray phase-contrast tomography to obtain an unprecedented direct 3D characterization of EAE lesions at micro-to-nano scales, with simultaneous imaging of the vascular and neuronal networks. We reveal EAE-mediated alterations down to the capillary network. Our findings shed light on how the disease and MSC treatment affect the tissues, and promote X-ray phase-contrast tomography as a powerful tool for studying neurovascular diseases and monitoring advanced therapies.