Phase retrieval for near-field X-ray imaging beyond linearisation or compact support

X-ray phase contrast imaging based on free space propagation relies on phase retrieval to obtain sharp images of micro- and nanoscale objects, with widespread applications in material science and biomedical research. For high resolution synchrotron experiments, phase retrieval is largely based on the single step reconstruction using the contrast transfer function approach (CTF), as introduced almost twenty years ago [Cloetens et al., Appl. Phys. Lett. 75, 2912 (1999)]. Notwithstanding its tremendous merits, this scheme makes stringent assumptions on the optical properties of the object, requiring, in particular, a weakly varying phase. In this work, we show how significant the loss in image quality becomes if these assumption are violated, and how phase retrieval can be easily improved by a simple scheme of alternating projections. Importantly, the approach demonstrated here uses the same input data and constraint sets as the conventional CTF-based phase retrieval, and is particularly well suited for the holographic regime

Phase-contrast tomography of sciatic nerves: image quality and experimental parameters

We present propagation-based phase-contrast tomography of mouse sciatic nerves stained with osmium, leading to an enhanced contrast in the myelin sheath around the axons, in order to visualize the threedimensional (3D) structure of the nerve. We compare different experimental parameters and show that contrast and resolution are high enough to identify single axons in the nerve, including characteristic functional structures such as Schmidt-Lanterman incisures

Three-dimensional mouse brain cytoarchitecture revealed by laboratory-based x-ray phase-contrast tomography

Studies of brain cytoarchitecture in mammals are routinely performed by serial sectioning of the specimen and staining of the sections. The procedure is labor-intensive and the 3D architecture can only be determined after aligning individual 2D sections, leading to a reconstructed volume with non-isotropic resolution. Propagation-based x-ray phase-contrast tomography offers a unique potential for high-resolution 3D imaging of intact biological specimen due to the high penetration depth and potential resolution. We here show that even compact laboratory CT at an optimized liquid-metal jet microfocus source combined with suitable phase-retrieval algorithms and a novel tissue preparation can provide cellular and subcellular resolution in millimeter sized samples of mouse brain. We removed water and lipids from entire mouse brains and measured the remaining dry tissue matrix in air, lowering absorption but increasing phase contrast. We present single-cell resolution images of mouse brain cytoarchitecture and show that axons can be revealed in myelinated fiber bundles. In contrast to optical 3D techniques our approach does neither require staining of cells nor tissue clearing, procedures that are increasingly difficult to apply with increasing sample and brain sizes. The approach thus opens a novel route for high-resolution high-throughput studies of brain architecture in mammals

X‐ray computed tomography and its potential in ecological research: A review of studies and optimization of specimen preparation

Imaging techniques are a cornerstone of contemporary biology. Over the last decades, advances in microscale imaging techniques have allowed fascinating new insights into cell and tissue morphology and internal anatomy of organisms across kingdoms. However, most studies so far provided snapshots of given reference taxa, describing organs and tissues under “idealized” conditions. Surprisingly, there is an almost complete lack of studies investigating how an organism′s internal morphology changes in response to environmental drivers. Consequently, ecology as a scientific discipline has so far almost neglected the possibilities arising from modern microscale imaging techniques. Here, we provide an overview of recent developments of X-ray computed tomography as an affordable, simple method of high spatial resolution, allowing insights into three-dimensional anatomy both in vivo and ex vivo. We review ecological studies using this technique to investigate the three-dimensional internal structure of organisms. In addition, we provide practical comparisons between different preparation techniques for maximum contrast and tissue differentiation. In particular, we consider the novel modality of phase contrast by self-interference of the X-ray wave behind an object (i.e., phase contrast by free space propagation). Using the cricket Acheta domesticus (L.) as model organism, we found that the combination of FAE fixative and iodine staining provided the best results across different tissues. The drying technique also affected contrast and prevented artifacts in specific cases. Overall, we found that for the interests of ecological studies, X-ray computed tomography is useful when the tissue or structure of interest has sufficient contrast that allows for an automatic or semiautomatic segmentation. In particular, we show that reconstruction schemes which exploit phase contrast can yield enhanced image quality. Combined with suitable specimen preparation and automated analysis, X-ray CT can therefore become a promising quantitative 3D imaging technique to study organisms′ responses to environmental drivers, in both ecology and evolution.</p

A phase-retrieval toolbox for X-ray holography and tomography

Propagation-based phase-contrast X-ray imaging is by now a well established imaging technique, which – as a full-field technique – is particularly useful for tomography applications. Since it can be implemented with synchrotron radiation and at laboratory micro-focus sources, it covers a wide range of applications. A limiting factor in its development has been the phase-retrieval step, which was often performed using methods with a limited regime of applicability, typically based on linearization. In this work, a much larger set of algorithms, which covers a wide range of cases (experimental parameters, objects and constraints), is compiled into a single toolbox – the HoloTomoToolbox – which is made publicly available. Importantly, the unified structure of the implemented phase-retrieval functions facilitates their use and performance test on different experimental data