Synchrotron radiation x-ray tomographic microscopy (SRXTM) of long bone

For each bone specimen, the bone was placed in a sample holder filled with cottons soaked in PBS to prevent the tissue dehydration and the displacement of the specimen during measurement. All SRXTM experiments were performed at the x-ray tomographic microscopy beamline (BL1.2W: XTM) of the Siam Photon Source (SPS), Synchrotron Light Research Institute (SLRI), Nakhon Ratchasima, Thailand. The image acquisition and parameters setup used in this experiment were based on the previous method of Tiyasatkulkovit and co-workers³⁰. All tomography datasets were collected with filtered polychromatic x-ray beam at the mean energy of 10.5 keV. The x-ray projections were acquired from the detection system comprising YAG-Ce scintillator coupled microscope (OptiquePeter, France) and PCO.edge 5.5 camera (PCO Imaging, Germany). This setup yielded an effective pixel size of 3.61 × 3.61 µm². The image processing and tomographic reconstruction was carried out by using Octopus Reconstruction software (Kohoutovice, Czech Republic).

As for the porosity measurement, the pore canals in the bone specimens were segmented and analyzed from the reconstructed slices by using Octopus Analysis software (Kohoutovice, Czech Republic). Herein, a fraction of the void volume over the analyzed volume (referred to as the volume of interest; VOI) were averaged from four regions in each specimen and expressed in terms of the percentage of porosity (% porosity—i.e., percent void volume). The 3D visualization and presentation of the volumetric data were rendered by using Drishti volume software (Canberra, Australia).