Analysis of microscopy techniques to measure segregation in continuous-cast steel slabs.

The accurate characterisation of centreline segregation requires precise measurements of composition variations over large length scales (10 - 1 $^{-1}$ m ${\rm {m}}$ ) across the centreline of the cast product, while having high resolution, sufficient to quantify the significant composition variations between dendrites due to microsegregation at very small length scales (10 - 5 m $^{-5}{\rm {m}}$ ). This study investigates the potential of a novel microscopy technique, named Synchrotron Micro X-ray Flurorescence (SMXRF), to generate large-scale high-resolution segregation maps from a steel sample taken from a thin slab caster. Two methods, Point Analysis and Regression Analysis, are proposed for SMXRF data calibration. By comparing with the traditional Laser-Induced Breakdown Spectroscopy (LIBS), and Electron Probe Micro Analyser (EPMA) techniques, we show that SMXRF is successful in quantitative characterisation of centreline segregation. Over large areas (e.g. 12 × $\times$ 16 mm 2 ${\rm {mm}}^2$ ) and at high resolution (10-50 µ m $\mu\text{m}$ pixel size) various techniques yield comparable outcomes in terms of composition maps and solute profiles. The findings also highlight the importance of both high spatial resolution and large field of view to have a quantitative, accurate, and efficient measurement tool to investigate segregation phenomena.

Comprehensive stereological assessment of the human lung using multiresolution computed tomography.

The application of stereology to lung casts and two-dimensional microscopy images is the gold standard for quantification of the human lung anatomy. However, these techniques are labor intensive, involving fixation, embedding, and histological sectioning of samples and thus have prevented comprehensive studies. Our objective was to demonstrate the application of stereology to volumetric multiresolution computed tomography (CT) to efficiently and extensively quantify the human lung anatomy. Nontransplantable donor lungs from individuals with no evidence of respiratory disease (n = 13) were air inflated, frozen at 10 cmH2O, and scanned using CT. Systematic uniform random samples were taken, scanned using micro-CT, and assessed using stereology. The application of stereology to volumetric CT imaging enabled comprehensive quantification of total lung volume, volume fractions of alveolar, alveolar duct, and tissue, mean linear intercept, alveolar surface area, alveolar surface area density, septal wall thickness, alveolar number, number-weighted mean alveolar volume, and the number and morphometry of terminal and transitional bronchioles. With the use of this data set, we found that women and men have the same number of terminal bronchioles (last generation of conducting airways), but men have longer terminal bronchioles, a smaller wall area percentage, and larger lungs due to a greater number of alveoli per acinus. The application of stereology to multiresolution CT imaging enables comprehensive analysis of the human lung parenchyma that identifies differences between men and women. The reported data set of normal donor lungs aged 25-77 yr provides reference data for future studies of chronic lung disease to determine exact changes in tissue pathology.NEW & NOTEWORTHY Stereology has been the gold standard to quantify the three-dimensional lung anatomy using two-dimensional microscopy images. However, such techniques are labor intensive. This study provides a method that applies stereology to volumetric computed tomography images of frozen whole human lungs and systematic uniform random samples. The method yielded a comprehensive data set on the small airways and parenchymal lung structures, highlighting morphometric sex differences and providing a reference data set for future pathological studies.

Nondestructive cryomicro-CT imaging enables structural and molecular analysis of human lung tissue.

Micro-computed tomography (CT) enables three-dimensional (3D) imaging of complex soft tissue structures, but current protocols used to achieve this goal preclude cellular and molecular phenotyping of the tissue. Here we describe a radiolucent cryostage that permits micro-CT imaging of unfixed frozen human lung samples at an isotropic voxel size of (11 µm)3 under conditions where the sample is maintained frozen at -30°C during imaging. The cryostage was tested for thermal stability to maintain samples frozen up to 8 h. This report describes the methods used to choose the materials required for cryostage construction and demonstrates that whole genome mRNA integrity and expression are not compromised by exposure to micro-CT radiation and that the tissue can be used for immunohistochemistry. The new cryostage provides a novel method enabling integration of 3D tissue structure with cellular and molecular analysis to facilitate the identification of molecular determinants of disease. NEW & NOTEWORTHY: The described micro-CT cryostage provides a novel way to study the three-dimensional lung structure preserved without the effects of fixatives while enabling subsequent studies of the cellular matrix composition and gene expression. This approach will, for the first time, enable researchers to study structural changes of lung tissues that occur with disease and correlate them with changes in gene or protein signatures.