Abstract
High-resolution episcopic microscopy (HREM) is a three-dimensional (3D) episcopic imaging modality based on the acquisition of two-dimensional (2D) images from the cut surface of a block of tissue embedded in resin. Such images, acquired serially through the entire length/depth of the tissue block, are aligned and stacked for 3D reconstruction. HREM has proven to be specifically advantageous when integrated in correlative multimodal imaging (CMI) pipelines. CMI creates a composite and zoomable view of exactly the same specimen and region of interest by (sequentially) correlating two or more modalities. CMI combines complementary modalities to gain holistic structural, functional, and chemical information of the entire sample and place molecular details into their overall spatiotemporal multiscale context. HREM has an advantage over in vivo 3D imaging techniques on account of better histomorphologic resolution while simultaneously providing volume data. HREM also has certain advantages over ex vivo light microscopy modalities. The latter can provide better cellular resolution but usually covers a limited area or volume of tissue, with limited 3D structural context. HREM has predominantly filled a niche in the phenotyping of embryos and characterisation of anatomic developmental abnormalities in various species. Under the umbrella of CMI, when combined with histopathology in a mutually complementary manner, HREM could find wider application in additional nonclinical and translational areas. HREM, being a modified histology technique, could also be incorporated into specialised preclinical pathology workflows. This review will highlight HREM as a versatile imaging platform in CMI approaches and present its benefits and limitations.
Highlights
High-resolution episcopic microscopy (HREM) can be combined with almost all upstream and downstream techniques in multimodal imaging pipelines. It proved to be the method of choice to correlatively reconstruct tumour capillaries and murine vasculature of sufficient contrast and quality at micrometre resolution in selected volumes of interest (VOIs), and to visualise minute anatomical structures and volume displays of mouse, chick, and zebrafish embryos in combination with optical coherence tomography (OCT), photoacoustic tomography (PAT), micro-US, micro-CT, and micro-MRI
We provide an exhaustive overview of published correlative multimodal imaging (CMI) workflows that integrated HREM to diagnose and characterise embryos or foetuses
To routinely implement CMI as an imaging procedure, several bottlenecks remain to be addressed: (1) it should be ensured that the handling of samples and preparation procedures are compatible across various modalities and that data quality is not compromised; (2) the same VOI must be located across imaging platforms with the help of softand hardware solutions; (3) software solutions that allow the correlation and handling of complex, multiscale, multimodal, and volumetric image data should be in place
Summary
CMI is capable of providing structural and functional information on a tissue sample that is visualised at different lateral resolutions and penetration depths across relevant scales. This essentially involves the application of two or more complementary modalities, which in combination, provide a more informative and composite view of normal and abnormal features of the tissue specimen [7,8,9]. (AFM) atomic force microscopy; (CT) computed tomography; (EM) electron microscopy; (HREM) high-resolution episcopic microscopy; (LSFM) light sheet fluorescence microscopy; (MPM) multiphoton microscopy; (MRI) magnetic resonance imaging; (MSI) mass spectrometry imaging; (OCT) optical coherence tomography; (OI) optical interferometry; (PAT).
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