X-ray-Based 3D Virtual Histology—Adding the Next Dimension to Histological Analysis

Abstract

Histology and immunohistochemistry of thin tissue sections have been the standard diagnostic procedure in many diseases for decades. This method is highly specific for particular tissue regions or cells, but mechanical sectioning of the specimens is required, which destroys the sample in the process and can lead to non-uniform tissue deformations. In addition, regions of interest cannot be located beforehand and the analysis is intrinsically two-dimensional. Micro X-ray computed tomography (μCT) on the other hand can provide 3D images at high resolution and allows for quantification of tissue structures, as well as the localization of small regions of interest. These advantages advocate the use of μCT for virtual histology tool with or without subsequent classical histology. This review summarizes the most recent examples of virtual histology and provides currently known possibilities of improving contrast and resolution of μCT. Following a background in μCT imaging, ex vivo staining procedures for contrast enhancement are presented as well as label-free virtual histology approaches and the technologies, which could rapidly advance it, such as phase-contrast CT. Novel approaches such as zoom tomography and nanoparticulate contrast agents will also be considered. The current evidence suggests that virtual histology may present a valuable addition to the workflow of histological analysis, potentially reducing the workload in pathology, refining tissue classification, and supporting the detection of small malignancies.

X-ray Micro-Computed Tomography for Nondestructive Three-Dimensional (3D) X-ray Histology

“…In contrast, FFPE histology specimens of soft tissues or demineralized hard tissues, with inherently low X-ray attenuation contrast between the tissue and the supporting matrix (paraffin wax), have previously been considered beyond the reach of routine μCT imaging. As noted above, specialized sample preparation protocols and X-ray systems can be used, 12 , 31 , 32 associated with several important disadvantages. For instance, X-ray contrast agents often lack binding-specific affinity for different tissue types and rely on the diffusion of heavy ions (ie, the contrast agent) into the tissue.…”

“…The latter is a slow process that requires immersion of the tissue into the ions' solution, which can take up to several days to complete. 32 Also, spatial and temporal anisotropy of stains' penetration can result in artificial contrast gradients between the core and the surface of the tissue and in stain-induced shrinkage of the tissue sample. 31 , 33 , 34 This complicates the interpretation, segmentation, and quantitative analysis of the microscopic tissue features of interest, such as epithelial surfaces, lymphatic vessels, or colonic crypt foci.…”

Correlative Light-Electron Microscopy (CLEM) and 3D Volume Imaging of Serial Block-Face Scanning Electron Microscopy (SBF-SEM) of Langerhans Islets

“…Correlative Light-Electron Microscopy (CLEM) and 3D Volume Imaging of Serial Block-Face… DOI: http://dx.doi.org /10.5772/intechopen.81716 in vivo imaging (IVIS @ ) containing micro/nano CT (computed tomography), has become an important tool for biomedical research (Figure 1) [25][26][27][28][29][30][31]. In particular, the development of a series of hybrid approaches in technological advancements of microscopy techniques, labelling tools, and fixation or preparation procedures allow correlating functional fluorescence microscopy data and ultrastructural information from a singular biological event.…”

High‐resolution 3D imaging of microvascular architecture in human glioma tissues using X‐ray phase‐contrast computed tomography as a potential adjunct to histopathology

“…Second, histology only provides two-dimensional (2D) information in a given cross section, 5 and regions of interest (ROIs) cannot be located beforehand, which may omit some important structures. 6 Although threedimensional (3D) volumes can be obtained by reconstruction of serial histological slices, this method has limitations relating to the 2D nature of histology and the deformation caused during processing. 7 3D analyzing of a whole specimen is crucial since the spatial maps of microvascular network cannot be fully presented in 2D representative sections.…”