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Advances in laboratory-based X-ray computed tomography (CT) have enabled X-ray 3D virtual histology. This method shows a great potential as a complementary technique to conventional 2D histology where extensive volumetric sampling is necessary. While formalin-fixed paraffin-embedded (FFPE) tissue blocks are the backbone of clinical histology, there exists no generic optimization, and technical study of the X-ray 3D virtual histology of FFPE blocks. X-ray micro-CT of FFPE blocks is studied and optimized in their native state within the cassette to minimize the interference of X-ray 3D virtual histology with clinical workflows and standards, hence facilitating the technology transfer to the clinics. The optimization is carried on the sample positioning, tungsten tubes acceleration voltage, and artifact reduction. Then propagation-based imaging of FFPE blocks is extensively discussed. Hierarchical (local) tomography and laminography are presented as viable approaches for achieving higher spatial resolutions. In the end, future perspectives are given by considering state-of-the-art micro-CT scanners using liquid-metal-jet sources, large-area detectors, and photon counting detectors. The results achieved here are generic and can be applicable to any laboratory-based scanner with a tungsten target source and cone-beam geometry. This article provides a starting point for anyone new to X-ray 3D virtual histology on FFPE blocks, but also serves as a useful source for more experienced users.INDEX TERMS 3D virtual histology, computed tomography, formalin-fixed paraffin-embedded blocks, phase-contrast imaging, propagation-based imaging, transport of intensity equation, X-ray imaging.
Advances in laboratory-based X-ray computed tomography (CT) have enabled X-ray 3D virtual histology. This method shows a great potential as a complementary technique to conventional 2D histology where extensive volumetric sampling is necessary. While formalin-fixed paraffin-embedded (FFPE) tissue blocks are the backbone of clinical histology, there exists no generic optimization, and technical study of the X-ray 3D virtual histology of FFPE blocks. X-ray micro-CT of FFPE blocks is studied and optimized in their native state within the cassette to minimize the interference of X-ray 3D virtual histology with clinical workflows and standards, hence facilitating the technology transfer to the clinics. The optimization is carried on the sample positioning, tungsten tubes acceleration voltage, and artifact reduction. Then propagation-based imaging of FFPE blocks is extensively discussed. Hierarchical (local) tomography and laminography are presented as viable approaches for achieving higher spatial resolutions. In the end, future perspectives are given by considering state-of-the-art micro-CT scanners using liquid-metal-jet sources, large-area detectors, and photon counting detectors. The results achieved here are generic and can be applicable to any laboratory-based scanner with a tungsten target source and cone-beam geometry. This article provides a starting point for anyone new to X-ray 3D virtual histology on FFPE blocks, but also serves as a useful source for more experienced users.INDEX TERMS 3D virtual histology, computed tomography, formalin-fixed paraffin-embedded blocks, phase-contrast imaging, propagation-based imaging, transport of intensity equation, X-ray imaging.
Malignant diseases are characterized by a critical trait known as invasiveness, where tumor cells tend to spread from the primary tissue layer into surrounding healthy tissues and distant organs. Presently, histopathology offers essential insights for diagnosing, classifying, predicting outcomes, and guiding patient-specific treatments. However, histology offers two-dimensional data from chosen cutting planes. Although 3D histological volumes can be generated through serial sectioning or whole slide imaging, this method is laborious, may introduce processing artefacts, and lacks isotropic spatial resolution. These limitations pose a considerable challenge to accurate diagnoses, particularly when dealing with micro-infiltrating carcinomas. These lesions, characterized by minute infiltrations, demand a three-dimensional representation for comprehensive visualization, essential for precise identification and assessment. Emerging X-ray-based virtual histology technology offers three-dimensional visualization of soft-tissue specimens, enabling virtual slicing in any direction or at any point. This approach can assist in guiding tissue sectioning for optimal representation of tumor cross sections during histological analysis. Micro-infiltrating carcinomas from the breast, cervix, and thyroid were imaged using X-ray phase-contrast microtomography (PhC-$$\mu$$ μ CT) at the Elettra synchrotron facility in Trieste, Italy. Comparative assessment of histological and CT slices by pathologists revealed that PhC-$$\mu$$ μ CT aids in classifying lesions by highlighting distinct tissue components and, notably, identifying tissue invasion. Reviewing a volume image allows pathologists to trace the entire lesion, identifying invasion sites that might be overlooked in individual or serial histological sections. Consequently, this proposed method could complement pathologists’ tools, potentially enhancing diagnoses by minimizing under-staging and reducing false negative results.
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