Imaging intact human organs with local resolution of cellular structures using hierarchical phase-contrast tomography

Abstract

Imaging intact human organs from the organ to the cellular scale in three dimensions is a goal of biomedical imaging. To meet this challenge, we developed hierarchical phase-contrast tomography (HiP-CT), an X-ray phase propagation technique using the European Synchrotron Radiation Facility (ESRF)’s Extremely Brilliant Source (EBS). The spatial coherence of the ESRF-EBS combined with our beamline equipment, sample preparation and scanning developments enabled us to perform non-destructive, three-dimensional (3D) scans with hierarchically increasing resolution at any location in whole human organs. We applied HiP-CT to image five intact human organ types: brain, lung, heart, kidney and spleen. HiP-CT provided a structural overview of each whole organ followed by multiple higher-resolution volumes of interest, capturing organotypic functional units and certain individual specialized cells within intact human organs. We demonstrate the potential applications of HiP-CT through quantification and morphometry of glomeruli in an intact human kidney and identification of regional changes in the tissue architecture in a lung from a deceased donor with coronavirus disease 2019 (COVID-19).

A multiscale X-ray phase-contrast tomography dataset of whole human left lung

“…Although synchrotron-based X-ray imaging can access finer anatomical detail than laboratory micro-CT 18,[20][21][22] , many bioimaging scenarios require further upscaling of the imaging throughput and accommodation of large sample size while maintaining microscopic resolution 23,24 . Thanks to the high X-ray photon flux and coherence achieved at modern fourth-generation synchrotron sources and careful design of the imaging protocol, it is now possible to image complete, large, partially dehydrated human organs in their entirety at micrometer resolution using hierarchical phase-contrast tomography (HiP-CT) 25 . It is a single-modality, multiscale imaging technique that employs propagation phase contrast from high-energy, polychromatic X-rays, flat-field correction, attenuation scanning protocol, along with efficient tomographic sampling and stitching pipeline to cover large, soft-tissue organs entirely.…”

“…The degassing procedure used a membrane pump to directly pump 32 above the PET sample jar with the lid open in a sealed vacuum glass dryer. Prior to imaging, the PET jar containing the lung, ethanol solution and agar embedding was placed in a custom-made sample holder to connect to the rotation stage at the synchrotron beamline 25 .…”

“…The implementation and capabilities of HiP-CT have been described in detail in a separate publication 25 . Here, we describe the settings used for lung imaging.…”

“…We used the half-acquisition (or half-object acquisition) 34 method developed at ESRF for imaging the VOIs at 6.5 µm and 2.5 µm in voxel size. For the entire lung, we developed a quarter-acquisition method 25 that includes the half-acquisition in combination with an annular scan to cover its complete horizontal extent (see Fig. 2).…”

“…2a,c): The incident X-ray energy averaged at ∼ 85 keV after filters, the propagation distance is 3475 mm. In total, two sets of 9990 projections were measured by the quarter-acquisition method 25 with an offset of 800 pixels for the half-acquisition. A step size of 2.2 mm in the vertical (z)…”