39 research outputs found

    Sample Preparation and Warping Accuracy for Correlative Multimodal Imaging in the Mouse Olfactory Bulb Using 2-Photon, Synchrotron X-Ray and Volume Electron Microscopy

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    Integrating physiology with structural insights of the same neuronal circuit provides a unique approach to understanding how the mammalian brain computes information. However, combining the techniques that provide both streams of data represents an experimental challenge. When studying glomerular column circuits in the mouse olfactory bulb, this approach involves e.g., recording the neuronal activity with in vivo 2-photon (2P) calcium imaging, retrieving the circuit structure with synchrotron X-ray computed tomography with propagation-based phase contrast (SXRT) and/or serial block-face scanning electron microscopy (SBEM) and correlating these datasets. Sample preparation and dataset correlation are two key bottlenecks in this correlative workflow. Here, we first quantify the occurrence of different artefacts when staining tissue slices with heavy metals to generate X-ray or electron contrast. We report improvements in the staining procedure, ultimately achieving perfect staining in ∌67% of the 0.6 mm thick olfactory bulb slices that were previously imaged in vivo with 2P. Secondly, we characterise the accuracy of the spatial correlation between functional and structural datasets. We demonstrate that direct, single-cell precise correlation between in vivo 2P and SXRT tissue volumes is possible and as reliable as correlating between 2P and SBEM. Altogether, these results pave the way for experiments that require retrieving physiology, circuit structure and synaptic signatures in targeted regions. These correlative function-structure studies will bring a more complete understanding of mammalian olfactory processing across spatial scales and time

    Functional and multiscale 3D structural investigation of brain tissue through correlative in vivo physiology, synchrotron microtomography and volume electron microscopy

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    Understanding the function of biological tissues requires a coordinated study of physiology and structure, exploring volumes that contain complete functional units at a detail that resolves the relevant features. Here, we introduce an approach to address this challenge: Mouse brain tissue sections containing a region where function was recorded using in vivo 2-photon calcium imaging were stained, dehydrated, resin-embedded and imaged with synchrotron X-ray computed tomography with propagation-based phase contrast (SXRT). SXRT provided context at subcellular detail, and could be followed by targeted acquisition of multiple volumes using serial block-face electron microscopy (SBEM). In the olfactory bulb, combining SXRT and SBEM enabled disambiguation of in vivo-assigned regions of interest. In the hippocampus, we found that superficial pyramidal neurons in CA1a displayed a larger density of spine apparati than deeper ones. Altogether, this approach can enable a functional and structural investigation of subcellular features in the context of cells and tissues

    Measuring cardiomyocyte cellular characteristics in cardiac hypertrophy using diffusion‐weighted MRI

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    Purpose: This paper presents a hierarchical modeling approach for estimating cardiomyocyte major and minor diameters and intracellular volume fraction (ICV) using diffusion‐weighted MRI (DWI) data in ex vivo mouse hearts. Methods: DWI data were acquired on two healthy controls and two hearts 3 weeks post transverse aortic constriction (TAC) using a bespoke diffusion scheme with multiple diffusion times ( Δ Δ \Delta ), q‐shells and diffusion encoding directions. Firstly, a bi‐exponential tensor model was fitted separately at each diffusion time to disentangle the dependence on diffusion times from diffusion weightings, that is, b‐values. The slow‐diffusing component was attributed to the restricted diffusion inside cardiomyocytes. ICV was then extrapolated at Δ = 0 Δ=0 \Delta =0 using linear regression. Secondly, given the secondary and the tertiary diffusion eigenvalue measurements for the slow‐diffusing component obtained at different diffusion times, major and minor diameters were estimated assuming a cylinder model with an elliptical cross‐section (ECS). High‐resolution three‐dimensional synchrotron X‐ray imaging (SRI) data from the same specimen was utilized to evaluate the biophysical parameters. Results: Estimated parameters using DWI data were (control 1/control 2 vs. TAC 1/TAC 2): major diameter—17.4 ÎŒ ÎŒ \mu m/18.0 ÎŒ ÎŒ \mu m versus 19.2 ÎŒ ÎŒ \mu m/19.0 ÎŒ ÎŒ \mu m; minor diameter—10.2 ÎŒ ÎŒ \mu m/9.4 ÎŒ ÎŒ \mu m versus 12.8 ÎŒ ÎŒ \mu m/13.4 ÎŒ ÎŒ \mu m; and ICV—62%/62% versus 68%/47%. These findings were consistent with SRI measurements. Conclusion: The proposed method allowed for accurate estimation of biophysical parameters suggesting cardiomyocyte diameters as sensitive biomarkers of hypertrophy in the heart

    Increased dose efficiency of breast CT with grating interferometry

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    Refraction-based x-ray imaging can overcome the fundamental contrast limit of computed tomography (CT), particularly in soft tissue, but so far has been constrained to high-dose ex vivo applications or required highly coherent x-ray sources, such as synchrotrons. Here we demonstrate that grating interferometry (GI) is more dose efficient than conventional CT in imaging of human breast under close-to-clinical conditions. Our system, based on a conventional source and commercial gratings, outperformed conventional CT for spatial resolutions better than 263 ”m and absorbed dose of 16 mGy. The sensitivity of GI is constrained by grating fabrication, and further progress will lead to significant improvements of clinical CT

    Zu den Wurzeln der Modernen Architektur, Teil I

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    Modern emerging technologies, such as additive manufacturing, bioprinting, and new material production, require novel metrology tools to probe fundamental high-speed dynamics happening in such systems. Here we demonstrate the application of the megahertz (MHz) European X-ray Free-Electron Laser (EuXFEL) to image the fast stochastic processes induced by a laser on water-filled capillaries with micrometer-scale spatial resolution. The EuXFEL provides superior contrast and spatial resolution compared to equivalent state-of-the-art synchrotron experiments. This work opens up new possibilities for the characterization of MHz stochastic processes on the nanosecond to microsecond time scales with object velocities up to a few kilometers per second using XFEL sources

    Validation of diffusion tensor MRI measurements of cardiac microstructure with structure tensor synchrotron radiation imaging.

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    Background Diffusion tensor imaging (DTI) is widely used to assess tissue microstructure non-invasively. Cardiac DTI enables inference of cell and sheetlet orientations, which are altered under pathological conditions. However, DTI is affected by many factors, therefore robust validation is critical. Existing histological validation is intrinsically flawed, since it requires further tissue processing leading to sample distortion, is routinely limited in field-of-view and requires reconstruction of three-dimensional volumes from two-dimensional images. In contrast, synchrotron radiation imaging (SRI) data enables imaging of the heart in 3D without further preparation following DTI. The objective of the study was to validate DTI measurements based on structure tensor analysis of SRI data. Methods One isolated, fixed rat heart was imaged ex vivo with DTI and X-ray phase contrast SRI, and reconstructed at 100 ÎŒm and 3.6 ÎŒm isotropic resolution respectively. Structure tensors were determined from the SRI data and registered to the DTI data. Results Excellent agreement in helix angles (HA) and transverse angles (TA) was observed between the DTI and structure tensor synchrotron radiation imaging (STSRI) data, where HADTI-STSRI = −1.4° ± 23.2° and TADTI-STSRI = −1.4° ± 35.0° (mean ± 1.96 standard deviation across all voxels in the left ventricle). STSRI confirmed that the primary eigenvector of the diffusion tensor corresponds with the cardiomyocyte long-axis across the whole myocardium. Conclusions We have used STSRI as a novel and high-resolution gold standard for the validation of DTI, allowing like-with-like comparison of three-dimensional tissue structures in the same intact heart free of distortion. This represents a critical step forward in independently verifying the structural basis and informing the interpretation of cardiac DTI data, thereby supporting the further development and adoption of DTI in structure-based electro-mechanical modelling and routine clinical applications

    X-ray phase-contrast imaging using near-field speckles

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    In the last decades, X-ray phase-contrast imaging has proven to be a powerful method for unveiling the inner structure of samples and is capable of visualising even minute density differences. Recently, speckle-based imaging (SBI), the youngest X-ray phase-sensitive technique, has received great interest due to its high sensitivity, quantitative character and relaxed requirements on the setup components and beam properties. This thesis is focussed on the development, experimental optimisation and applications of SBI, with the aim of simplifying its implementation, increasing its flexibility and expanding its potential. For this, a robust, flexible data acquisition and reconstruction approach, the unified modulated pattern analysis (UMPA), was developed, which lifts previous constraints of SBI. UMPA allows for tuning of the sensitivity and spatial resolution by adjusting the scan and reconstruction parameters. It is applicable not only to random speckle, but also periodic interference patterns, bridging the gap and improving the performance of both speckle- and single-grating-based techniques. Following the first demonstration of UMPA, its potential for a range of applications is illustrated in this thesis. It is shown that UMPA can be employed for X-ray optics characterisation to quantify aberrations in the focussing behaviour of X-ray refractive lenses with high precision and accuracy. UMPA phase tomography is applied to the field of biomedical imaging for high-sensitivity three-dimensional (3D) virtual histology of unstained, hydrated soft tissue, giving unprecedented structural and quantitative density information. Further developments of SBI explored in this thesis include the testing of novel customisable speckle diffusers, the extension of SBI to higher X-ray energies for geology and materials science applications, and the demonstration of UMPA at a laboratory X-ray source. These progresses promise new possibilities of SBI for high-sensitivity, robust and high-throughput imaging in previously inaccessible fields and make SBI accessible to a wider range of users in research and industry

    State of the art of X-ray speckle-based phase-contrast and dark-field imaging

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    In the past few years, X-ray phase-contrast and dark-field imaging have evolved to be invaluable tools for non-destructive sample visualisation, delivering information inaccessible by conventional absorption imaging. X-ray phase-sensing techniques are furthermore increasingly used for at-wavelength metrology and optics characterisation. One of the latest additions to the group of differential phase-contrast methods is the X-ray speckle-based technique. It has drawn significant attention due to its simple and flexible experimental arrangement, cost-effectiveness and multimodal character, amongst others. Since its first demonstration at highly brilliant synchrotron sources, the method has seen rapid development, including the translation to polychromatic laboratory sources and extension to higher-energy X-rays. Recently, different advanced acquisition schemes have been proposed to tackle some of the main limitations of previous implementations. Current applications of the speckle-based method range from optics characterisation and wavefront measurement to biomedical imaging and materials science. This review provides an overview of the state of the art of the X-ray speckle-based technique. Its basic principles and different experimental implementations as well as the the latest advances and applications are illustrated. In the end, an outlook for anticipated future developments of this promising technique is given.</p

    Simulations of multi-contrast X-ray imaging using near-field speckles

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    X-ray dark-field and phase-contrast imaging using near-field speckles is a novel technique that overcomes limitations inherent in conventional absorption X-ray imaging, i.e. poor contrast for features with similar density. Speckle-based imaging yields a wealth of information with a simple setup tolerant to polychromatic and divergent beams, and simple data acquisition and analysis procedures. Here, we present a simulation software used to model the image formation with the speckle-based technique, and we compare simulated results on a phantom sample with experimental synchrotron data. Thorough simulation of a speckle-based imaging experiment will help for better understanding and optimising the technique itself

    Simulations of x-ray speckle-based dark-field and phase-contrast imaging with a polychromatic beam

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    Following the first experimental demonstration of x-ray speckle-based multimodal imaging using a polychromatic beam [I. Zanette et al., Phys. Rev. Lett. 112(25), 253903 (2014)], we present a simulation study on the effects of a polychromatic x-ray spectrum on the performance of this technique. We observe that the contrast of the near-field speckles is only mildly influenced by the bandwidth of the energy spectrum. Moreover, using a homogeneous object with simple geometry, we characterize the beam hardening artifacts in the reconstructed transmission and refraction angle images, and we describe how the beam hardening also affects the dark-field signal provided by speckle tracking. This study is particularly important for further implementations and developments of coherent speckle-based techniques at laboratory x-ray sources
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