Project description:Micro-CT provides 3D volume imaging with spatial resolution at the micrometre scale. We investigated the optimal human placenta tissue preparation (contrast agent, perfusion pressure, perfusion location and perfusion vessel) and imaging (energy, target material, exposure time and frames) parameters. Microfil (Flow Tech, Carver, MA) produced better fill than Barium sulphate (84.1%(±11.5%)vs70.4%(±18.02%) p = 0.01). Perfusion via umbilical artery produced better fill than via chorionic vessels (83.8%(±17.7%)vs78.0%(±21.9%), p < 0.05), or via umbilical vein (83.8%(±16.4%)vs69.8%(±20.3%), p < 0.01). Imaging at 50 keV with a molybdenum target produced the best contrast to noise ratio. We propose this method to enable quantification and comparison of the human fetoplacental vascular tree.
Project description:Low inherent contrast in soft tissues has been limiting the use of X-ray absorption micro-computed tomography (micro-CT) to access high-resolution structural information of animal organs. The staining agents used in micro-CT to improve the contrast fail in providing high-quality images of whole organs of animals due to diffusion problems of the staining agent into the sample. We demonstrate a staining protocol that incorporates a biochemical conditioning step prior to exposure to the staining agent that succeeds in overcoming the diffusion problems, thus quickly providing high-quality micro-CT images of whole organs of mammals. Besides of yielding non-distorted three-dimensional information at the same spatial resolution accessible in histological sections, micro-CT images of whole organs stained by our method enable easy screening of slices along any direction of the volume thus demonstrating new possibilities of structural analysis in biomedical science.
Project description:Characterisation and quantification of tissue structures is limited by sectioning-induced artefacts and by the difficulties of visualising and segmenting 3D volumes. Here we demonstrate that, even in the absence of X-ray contrast agents, X-ray computed microtomography (microCT) and nanotomography (nanoCT) can circumvent these problems by rapidly resolving compositionally discrete 3D tissue regions (such as the collagen-rich adventitia and elastin-rich lamellae in intact rat arteries) which in turn can be segmented due to their different X-ray opacities and morphologies. We then establish, using X-ray tomograms of both unpressurised and pressurised arteries that intra-luminal pressure not only increases lumen cross-sectional area and straightens medial elastic lamellae but also induces profound remodelling of the adventitial layer. Finally we apply microCT to another human organ (skin) to visualise the cell-rich epidermis and extracellular matrix-rich dermis and to show that conventional histological and immunohistochemical staining protocols are compatible with prior X-ray exposure. As a consequence we suggest that microCT could be combined with optical microscopy to characterise the 3D structure and composition of archival paraffin embedded biological materials and of mechanically stressed dynamic tissues such as the heart, lungs and tendons.
Project description:The number of the Asbestos Bodies (AB), i.e. asbestos that developed an iron-protein coating during its permanence in biological tissues, is one of the most accessible markers of asbestos exposure in individuals. The approaches developed to perform AB count in biological tissues are based on the manual examination of tissue digests or histological sections by means of light or electron microscopies. Although these approaches are well established and relatively accessible, manual examination is time-consuming and can be reader-dependent. Besides, approximations are applied because of the limitations of 2D readings and to speed up manual counts. In addition, sample preparation using tissue digests require an amount of tissue that can only be obtained by invasive surgery or post-mortem sampling. In this paper, we propose a new approach to AB counting based on non-destructive 3D imaging, which has the potential to overcome most of the limitations of conventional approaches. This method allows automating the AB count and determining their morphometry distribution in bulk tissue samples (ideally non-invasive needle biopsies), with minimal sample preparation and avoiding approximations. Although the results are promising, additional testing on a larger number of AB-containing biological samples would be required to fully validate the method.
Project description:Historically, micro-computed tomography (μCT) has been considered unsuitable for histologic analysis of unstained formalin-fixed, paraffin-embedded soft tissue biopsy specimens because of a lack of image contrast between the tissue and the paraffin. However, we recently demonstrated that μCT can successfully resolve microstructural detail in routinely prepared tissue specimens. Herein, we illustrate how μCT imaging of standard formalin-fixed, paraffin-embedded biopsy specimens can be seamlessly integrated into conventional histology workflows, enabling nondestructive three-dimensional (3D) X-ray histology, the use and benefits of which we showcase for the exemplar of human lung biopsy specimens. This technology advancement was achieved through manufacturing a first-of-kind μCT scanner for X-ray histology and developing optimized imaging protocols, which do not require any additional sample preparation. 3D X-ray histology allows for nondestructive 3D imaging of tissue microstructure, resolving structural connectivity and heterogeneity of complex tissue networks, such as the vascular network or the respiratory tract. We also demonstrate that 3D X-ray histology can yield consistent and reproducible image quality, enabling quantitative assessment of a tissue's 3D microstructures, which is inaccessible to conventional two-dimensional histology. Being nondestructive, the technique does not interfere with histology workflows, permitting subsequent tissue characterization by means of conventional light microscopy-based histology, immunohistochemistry, and immunofluorescence. 3D X-ray histology can be readily applied to a plethora of archival materials, yielding unprecedented opportunities in diagnosis and research of disease.
Project description:Comparative anatomy is an important tool for investigating evolutionary relationships among species, but the lack of scalable imaging tools and stains for rapidly mapping the microscale anatomies of related species poses a major impediment to using comparative anatomy approaches for identifying evolutionary adaptations. We describe a method using synchrotron source micro-x-ray computed tomography (syn-μXCT) combined with machine learning algorithms for high-throughput imaging of Lepidoptera (i.e., butterfly and moth) eyes. Our pipeline allows for imaging at rates of ~15 min/mm3 at 600 nm3 resolution. Image contrast is generated using standard electron microscopy labeling approaches (e.g., osmium tetroxide) that unbiasedly labels all cellular membranes in a species-independent manner thus removing any barrier to imaging any species of interest. To demonstrate the power of the method, we analyzed the 3D morphologies of butterfly crystalline cones, a part of the visual system associated with acuity and sensitivity and found significant variation within six butterfly individuals. Despite this variation, a classic measure of optimization, the ratio of interommatidial angle to resolving power of ommatidia, largely agrees with early work on eye geometry across species. We show that this method can successfully be used to determine compound eye organization and crystalline cone morphology. Our novel pipeline provides for fast, scalable visualization and analysis of eye anatomies that can be applied to any arthropod species, enabling new questions about evolutionary adaptations of compound eyes and beyond.
Project description:Laboratory x-ray micro-computed tomography (micro-CT) is a fast-growing method in scientific research applications that allows for non-destructive imaging of morphological structures. This paper provides an easily operated "how to" guide for new potential users and describes the various steps required for successful planning of research projects that involve micro-CT. Background information on micro-CT is provided, followed by relevant setup, scanning, reconstructing, and visualization methods and considerations. Throughout the guide, a Jackson's chameleon specimen, which was scanned at different settings, is used as an interactive example. The ultimate aim of this paper is make new users familiar with the concepts and applications of micro-CT in an attempt to promote its use in future scientific studies.
Project description:As a novel low-side-effect cancer therapy, photo-immunotherapy (PIT) is based on conjugating monoclonal antibody (mAb) with a near-infrared (NIR) phthalocyanine dye IRDye700DX (IR 700). IR700 is not only fluorescent to be used as an imaging agent, but also phototoxic. When illuminating with NIR light, PIT can induce highly-selective cancer cell death while leaving most of tumor blood vessels unharmed, leading to an effect termed super-enhanced permeability and retention (SUPR), which can significantly improve the effectiveness of anti-cancer drug. Currently, the therapeutic effects of PIT are monitored using 2D macroscopic fluorescence reflectance imager, which lacks the resolution and depth information to reveal the 3D distribution of mAb-IR700. In the study, we applied a multi-modal optical imaging approach including high-resolution optical coherence tomography (OCT) and high-sensitivity fluorescence laminar optical tomography (FLOT), to provide 3D tumor micro-structure and micro-distribution of mAb-IR700 in the tumor simultaneously during PIT in situ and in vivo. The multi-wavelength FLOT can also provide the blood vessels morphology of the tumor. Thus, the 3D FLOT reconstructed images allow us to evaluate the IR700 fluorescence distribution change with respect to the blood vessels and at different tumor locations/depths non-invasively, thereby enabling evaluation of the therapeutic effects in vivo and optimization of treatment regimens accordingly. The mAb-IR700 can access more tumor areas after PIT treatment, which can be explained by increased vascular permeability immediately after NIR-PIT. Two-photon microscopy was also used to record the mAb-IR700 on the tumor surface near the blood vessels to verify the results.
Project description:Bone quality is commonly used to diagnose bone diseases such as osteoporosis, with many studies focusing on microarchitecture for fracture prediction. In this study a bovine distal femur was imaged using both micro-computed tomography (µCT) and tomosynthesis using focal construct geometry (FCG) for comparison of microarchitectural parameters. Six regions of interest (ROIs) were compared between the two imaging modalities, with both global and adaptive methods used to binarize the images. FCG images were downsampled to the same pixel size as the µCT images. Bone morphometrics were determined using BoneJ, for each imaging modality, binarization technique and ROI. Bone area/total area was found to have few significant differences between FCG and µCT (p < 0.05 for two of six ROIs). Fractal Dimension had only one significant difference (p < 0.05 for one of six ROIs) between µCT and downsampled FCG (where pixel size was equalized). Trabecular thickness and trabecular spacing were observed to follow trends as observed for the corresponding µCT images, although many absolute values were significantly different (p < 0.05 for between one and six ROIs depending on image types used). This study demonstrates the utility of tomosynthesis for measurement of microarchitectural morphometrics.