Fabrication of a fractal pattern device for focus characterizations of X-ray imaging systems by Si deep reactive ion etching and bottom-up Au electroplating.

Precisely aligned optical components are crucial prerequisites for X-ray tomography at high resolution. We propose a device with a fractal pattern for precise automatic focusing. The device is etched in a Si substrate by deep reactive ion etching and then filled by a self-terminating bottom-up Au electroplating process. The fractal nature of the device produces an X-ray transmission image with globally homogeneous macroscopic visibility and high local contrast for pixel sizes in the range of 0.165 µm to 11 µm, while the high absorption contrast provided between Au and Si enables its use for X-ray energies ranging from 12 keV to 40 keV.

High sensitivity X-ray phase contrast imaging by laboratory grating-based interferometry at high Talbot order geometry.

X-ray phase contrast imaging is a powerful analysis technique for materials science and biomedicine. Here, we report on laboratory grating-based X-ray interferometry employing a microfocus X-ray source and a high Talbot order (35th) asymmetric geometry to achieve high angular sensitivity and high spatial resolution X-ray phase contrast imaging in a compact system (total length <1 m). The detection of very small refractive angles (∼50 nrad) at an interferometer design energy of 19 keV was enabled by combining small period X-ray gratings (1.0, 1.5 and 3.0 µm) and a single-photon counting X-ray detector (75 µm pixel size). The performance of the X-ray interferometer was fully characterized in terms of angular sensitivity and spatial resolution. Finally, the potential of laboratory X-ray phase contrast for biomedical imaging is demonstrated by obtaining high resolution X-ray phase tomographies of a mouse embryo embedded in solid paraffin and a formalin-fixed full-thickness sample of human left ventricle in water with a spatial resolution of 21.5 µm.

Laboratory X-ray interferometry imaging with a fan-shaped source grating.

The orientation mismatch between the cone beam of an X-ray tube and the grating lines in a flat substrate remains a big challenge for laboratory grating-based X-ray interferometry, since it severely limits the imaging field of view. Here, we fabricated fan-shaped G0 source gratings by modulating the electric field during the deep reactive ion etching of silicon. The gold electroplated fan-shaped G0 grating (3.0 µm pitch) in a 20 keV interferometer improves the uniformity of the field of view with an increase of average visibility from 16.2% to 18.5% and a better angular sensitivity (by a factor 5.8) at the edges.

Correction to: Towards clinical grating-interferometry mammography.

The article Towards clinical grating-interferometry mammography, written by Carolina Arboleda, Zhentian Wang, Konstantins Jefimovs, Thomas Koehler, Udo Van Stevendaal, Norbert Kuhn, Bernd David, Sven Prevrhal, Kristina Lång, Serafino Forte, Rahel Antonia Kubik-Huch, Cornelia Leo.

Towards clinical grating-interferometry mammography.

OBJECTIVES: Grating-interferometry-based mammography (GIM) might facilitate breast cancer detection, as several research works have demonstrated in a pre-clinical setting, since it is able to provide attenuation, differential phase contrast, and scattering images simultaneously. In order to translate this technique to the clinics, it has to be adapted to cover a large field-of-view within a clinically acceptable exposure time and radiation dose. METHODS: We set up a grating interferometer that fits into a standard mammography system and fulfilled the aforementioned conditions. Here, we present the first mastectomy images acquired with this experimental device. RESULTS AND CONCLUSION: Our system performs at a mean glandular dose of 1.6 mGy for a 5-cm-thick, 18%-dense breast, and a field-of-view of 26 × 21 cm2. It seems to be well-suited as basis for a clinical-environment device. Further, dark-field signals seem to support an improved lesion visualization. Evidently, the effective impact of such indications must be evaluated and quantified within the context of a proper reader study. KEY POINTS: • Grating-interferometry-based mammography (GIM) might facilitate breast cancer detection, since it is sensitive to refraction and scattering and thus provides additional tissue information. • The most straightforward way to do grating-interferometry in the clinics is to modify a standard mammography device. • In a first approximation, the doses given with this technique seem to be similar to those of conventional mammography.

Visibility simulation of realistic grating interferometers including grating geometries and energy spectra.

The performance of X-ray and neutron grating interferometers is characterised by their visibility, which is a measure for the maximum achievable contrast. In this study we show how the real grating geometry in a grating interferometer with three gratings impacts the interference and self projection that leads to visibility in the first place. We quantify the individual contributions of wavelength distributions and grating shapes in terms of visibility reduction by determining the absolute as well as relative effect of each contribution. The understanding of the impact of changed geometry and wavelength distributions on the interference of neutrons/X-rays allows us to present the first fully quantitative model of a grating interferometer setup. We demonstrate the capability of the simulation framework by building a model of the neutron grating interferometer at the ICON beamline and directly comparing simulated and measured visibility values. The general nature of the model makes it possible to extend it to any given grating interferometer for both X-rays and neutrons.

Interpretation and Utility of the Moments of Small-Angle X-Ray Scattering Distributions.

Small angle x-ray scattering has been proven to be a valuable method for accessing structural information below the spatial resolution limit implied by direct imaging. Here, we theoretically derive the relation that links the subpixel differential phase signal provided by the sample to the moments of scattering distributions accessible by refraction sensitive x-ray imaging techniques. As an important special case we explain the scatter or dark-field contrast in terms of the sample's phase signal. Further, we establish that, for binary phase objects, the nth moment scales with the difference of the refractive index decrement to the power of n. Finally, we experimentally demonstrate the utility of the moments by quantitatively determining the particle sizes of a range of powders with a laboratory-based setup.

2D-Omnidirectional Hard-X-Ray Scattering Sensitivity in a Single Shot.

X-ray scattering imaging can provide complementary information to conventional absorption based radiographic imaging about the unresolved microstructures of a sample. The scattering signal can be accessed with various methods based on coherent illumination, which span from self-imaging to speckle scanning. The directional sensitivity of the existing real space imaging methods is limited to a few directions on the imaging plane and requires scanning of the optical components, or the rotation of either the sample or the imaging setup, in order to cover the full range of possible scattering directions. In this Letter the authors propose a new method that allows the simultaneous acquisition of scattering images in all possible directions in a single shot. This is achieved by a specialized phase grating and a detector with sufficient spatial resolution to record the generated interference fringe. The structural length scale sensitivity of the system can be tuned by varying its geometry for a fixed grating design. Taking into account ongoing developments in the field of compact x-ray sources that allow high brightness and sufficient spatial coherence, the applicability of omnidirectional scattering imaging in industrial and medical settings is boosted significantly.

Anisotropic de Gennes Narrowing in Confined Fluids.

The collective diffusion of dense fluids in spatial confinement is studied by combining high-energy (21 keV) x-ray photon correlation spectroscopy and small-angle x-ray scattering from colloid-filled microfluidic channels. We find the structural relaxation in confinement to be slower compared to the bulk. The collective dynamics is wave vector dependent, akin to the de Gennes narrowing typically observed in bulk fluids. However, in stark contrast to the bulk, the structure factor and de Gennes narrowing in confinement are anisotropic. These experimental observations are essential in order to develop a microscopic theoretical description of collective diffusion of dense fluids in confined geometries.

Mass spectrometry-based metabolomics of single yeast cells.

Single-cell level measurements are necessary to characterize the intrinsic biological variability in a population of cells. In this study, we demonstrate that, with the microarrays for mass spectrometry platform, we are able to observe this variability. We monitor environmentally (2-deoxy-D-glucose) and genetically (ΔPFK2) perturbed Saccharomyces cerevisiae cells at the single-cell, few-cell, and population levels. Correlation plots between metabolites from the glycolytic pathway, as well as with the observed ATP/ADP ratio as a measure of cellular energy charge, give biological insight that is not accessible from population-level metabolomic data.

Resolution pattern for mass spectrometry imaging.

RATIONALE: Up to now, there is no 'gold standard' for determining the resolution of a mass spectrometry imaging (MSI) setup (comprising the instrument, the sample preparation, the sample and the instrument settings). A standard sample in combination with a standard protocol to define the MSI resolution would be desirable in order to compare the setups of different laboratories, and as a regular quality control/performance check. METHODS: Microstructured resolution patterns were fabricated that can be used to determine the spatial resolution in MSI experiments, down to the range of a few µm. Two different strategies were employed, one where the resolution pattern is laser machined into a thin metal foil, which can be placed over a sample to be imaged, and a second one where hydrophilic grooves are machined into an omniphobic coating covering the surface of an indium tin oxide covered glass slide. When dragging a sample solution over the slide's surface, the sample is automatically retained in the hydrophilic grooves, but repelled by the omniphobic coating. RESULTS: The technology was tested on a commercial matrix-assisted laser desorption/ionization (MALDI) imaging instrument, and a spatial resolution in the vicinity of 50 µm was determined. The finest features of the microstructured resolution patterns are compatible with the best spatial resolution of MALDI imaging systems available to date. CONCLUSIONS: The use of metal resolution grids or glass slides with hydrophilic/hydrophobic structures is suitable for the convenient determination of the resolution limit of the MALDI imaging instrument as determined by its hardware. These structures are straightforward both to produce and to use.

High-resolution droplet-based fractionation of nano-LC separations onto microarrays for MALDI-MS analysis.

We present a robust droplet-based device, which enables the fractionation of ultralow flow rate nanoflow liquid chromatography (nano-LC) eluate streams at high frequencies and high peak resolution. This is achieved by directly interfacing the separation column to a micro T-junction, where the eluate stream is compartmentalized into picoliter droplets. This immediate compartmentalization prevents peak dispersion during eluate transport and conserves the chromatographic performance. Subsequently, nanoliter eluate fractions are collected at a rate of one fraction per second on a high-density microarray to retain the separation with high temporal resolution. Chromatographic separations of up to 45 min runtime can thus be archived on a single microarray possessing 2700 sample spots. The performance of this device is demonstrated by fractionating the separation of a tryptic digest of a known protein mixture onto the microarray chip and subsequently analyzing the sample archive using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). Resulting peak widths are found to be significantly reduced compared to standard continuous flow spotting technologies as well as in comparison to a conventional nano-LC-electrospray ionization-mass spectrometry interface. Moreover, we demonstrate the advantage of our high-definition nanofractionation device by applying two different MALDI matrices to all collected fractions in an alternating fashion. Since the information that is obtained from a MALDI-MS measurement depends on the choice of MALDI matrix, we can extract complementary information from neighboring spots containing almost identical composition but different matrices.

Molecular phenotypic profiling of a Saccharomyces cerevisiae strain at the single-cell level.

Studying cell-to-cell heterogeneity requires techniques which robustly deliver reproducible results with single-cell sensitivity. Through a new fabrication method for the microarrays for mass spectrometry (MAMS) platform, we now have attained robustness and reproducibility in our single-cell level mass spectrometry measurements that allowed us to combine single-cell MAMS-based measurements from different days and samples. By combining multiple measurements, we were able to identify three co-existing phenotypes in an isogenic population of Saccharomyces cerevisiae characterized by distinctively different levels of glycolytic intermediates.

Implementation of a dual-phase grating interferometer for multi-scale characterization of building materials by tunable dark-field imaging.

The multi-scale characterization of building materials is necessary to understand complex mechanical processes, with the goal of developing new more sustainable materials. To that end, imaging methods are often used in materials science to characterize the microscale. However, these methods compromise the volume of interest to achieve a higher resolution. Dark-field (DF) contrast imaging is being investigated to characterize building materials in length scales smaller than the resolution of the imaging system, allowing a direct comparison of features in the nano-scale range and overcoming the scale limitations of the established characterization methods. This work extends the implementation of a dual-phase X-ray grating interferometer (DP-XGI) for DF imaging in a lab-based setup. The interferometer was developed to operate at two different design energies of 22.0 keV and 40.8 keV and was designed to characterize nanoscale-size features in millimeter-sized material samples. The good performance of the interferometer in the low energy range (LER) is demonstrated by the DF retrieval of natural wood samples. In addition, a high energy range (HER) configuration is proposed, resulting in higher mean visibility and good sensitivity over a wider range of correlation lengths in the nanoscale range. Its potential for the characterization of mineral building materials is illustrated by the DF imaging of a Ketton limestone. Additionally, the capability of the DP-XGI to differentiate features in the nanoscale range is proven with the dark-field of Silica nanoparticles at different correlation lengths of calibrated sizes of 106 nm, 261 nm, and 507 nm.

Interfacing droplet microfluidics with matrix-assisted laser desorption/ionization mass spectrometry: label-free content analysis of single droplets.

Droplet-based microfluidic systems have become a very powerful tool to miniaturize chemical and biological reactions. However, droplet content analysis remains challenging and relies almost exclusively on optical methods such as fluorescence spectroscopy. Hence, labeling of the analyte is typically required which impedes a more universal applicability of microdroplets. Here we present a novel interface coupling droplet microfluidics and matrix-assisted laser desorption/ionization (MALDI) mass spectrometry for label-free content analysis of single droplets. Nanoliter aqueous droplets immersed in perfluorinated oil are created in a microfluidic T-junction, transferred into a capillary, and deposited on a high-density microarray MALDI plate mounted on a motorized xy-stage. The fully automated system is robust and reliable due to two unique features. First, a simple optical droplet detection system is used to synchronize stage movement and exit of droplets from the capillary. Second, the microarray plate contains an array of over 26,000 hydrophilic spots within a hydrophobic coating, each spot acting as a recipient to confine the droplets and to prevent cross-contamination. The MALDI matrix can also be applied using our system by spotting matrix droplets on the microarray in a separate run. To demonstrate the potential of our system, we studied the enzymatic cleavage of angiotensin I by angiotensin converting enzyme and monitored the increasing concentration of the product angiotensin II over time. The interface provides a robust and fully automated method for rapid label-free and information-rich content analysis of single droplets. With the high number of droplets per plate, this method is particularly suitable for high-throughput screening applications.

Self-aliquoting microarray plates for accurate quantitative matrix-assisted laser desorption/ionization mass spectrometry.

Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is a fast analysis tool employed for the detection of a broad range of analytes. However, MALDI-MS has a reputation of not being suitable for quantitative analysis. Inhomogeneous analyte/matrix co-crystallization, spot-to-spot inhomogeneity, as well as a typically low number of replicates are the main contributing factors. Here, we present a novel MALDI sample target for quantitative MALDI-MS applications, which addresses the limitations mentioned above. The platform is based on the recently developed microarray for mass spectrometry (MAMS) technology and contains parallel lanes of hydrophilic reservoirs. Samples are not pipetted manually but deposited by dragging one or several sample droplets with a metal sliding device along these lanes. Sample is rapidly and automatically aliquoted into the sample spots due to the interplay of hydrophilic/hydrophobic interactions. With a few microliters of sample, it is possible to aliquot up to 40 replicates within seconds, each aliquot containing just 10 nL. The analyte droplet dries immediately and homogeneously, and consumption of the whole spot during MALDI-MS analysis is typically accomplished within few seconds. We evaluated these sample targets with respect to their suitability for use with different samples and matrices. Furthermore, we tested their application for generating calibration curves of standard peptides with α-cyano-4-hdydroxycinnamic acid as a matrix. For angiotensin II and [Glu(1)]-fibrinopeptide B we achieved coefficients of determination (r(2)) greater than 0.99 without the use of internal standards.

Analysis of single algal cells by combining mass spectrometry with Raman and fluorescence mapping.

In order to investigate metabolic properties of single cells of freshwater algae (Haematococcus pluvialis), we implement matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) in combination with microspectroscopic mapping. Straightforward coupling of these two detection platforms was possible thanks to the self-aliquoting properties of micro-arrays for mass spectrometry (MAMS). Following Raman and fluorescence imaging, the isolated cells were covered with a MALDI matrix for targeted metabolic analysis by MALDI-MS. The three consecutive measurements carried out on the same cells yielded complementary information. Using this method, we were able to study the encystment of H. pluvialis - by monitoring the adenosine triphosphate (ATP) to adenosine diphosphate (ADP) ratio during the build-up of astaxanthin in the cells as well as the release of ß-carotene, the precursor of astaxanthin, into the cytosol.

Helium focused ion beam fabricated plasmonic antennas with sub-5 nm gaps.

We demonstrate a reliable fabrication method to produce plasmonic dipole nanoantennas with gap values in the range of 3.5-20 nm. The method combines electron beam lithography to create gold nanorods and helium focused ion beam milling to cut the gaps. Results show a reproducibility within 1 nm. Scattering spectra of antennas show a red shift of resonance wavelengths and an increase of the intensity of resonance peaks with a decrease of the gap size, which is in agreement with finite element simulations. The measured refractive index sensitivity was about 250 nm per refractive index unit for antennas with gap values below 5 nm.

Editorial for the Special Issue on Recent Advances in Reactive Ion Etching and Applications of High-Aspect-Ratio Microfabrication.

Reactive ion etching (RIE) is the dominating technology for micromachining semiconductors with a high aspect ratio (HAR) [...].

The choice of an autocorrelation length in dark-field lung imaging.

Respiratory diseases are one of the most common causes of death, and their early detection is crucial for prompt treatment. X-ray dark-field radiography (XDFR) is a promising tool to image objects with unresolved micro-structures such as lungs. Using Talbot-Lau XDFR, we imaged inflated porcine lungs together with Polymethylmethacrylat (PMMA) microspheres (in air) of diameter sizes between 20 and 500 [Formula: see text] over an autocorrelation range of 0.8-5.2 [Formula: see text]. The results indicate that the dark-field extinction coefficient of porcine lungs is similar to that of densely-packed PMMA spheres with diameter of [Formula: see text], which is approximately the mean alveolar structure size. We evaluated that, in our case, the autocorrelation length would have to be limited to [Formula: see text] in order to image [Formula: see text]-thick lung tissue without critical visibility reduction (signal saturation). We identify the autocorrelation length to be the critical parameter of an interferometer that allows to avoid signal saturation in clinical lung dark-field imaging.