EUROPEAN CONGRESS AND EXHIBITION ON ADVANCED MATERIALS AND PROCESSES - EUROMAT 2021

Bone is a biological nanocomposite, combining toughness and strength with a low weight. Similar to other complex composites, the hierarchical organization of mineralized tissues affects drastically their mechanical performance across different scales. The elementary building block of bone at the microscale consists of mineralized collagen fibril (MCF, each 10-300 nm in diameter) arrays embedded in an extrafibrillar matrix. Human cortical bone exhibits complex structural arrangement of MCF that may be affected by metabolic bone conditions like osteoporosis or Paget’s disease.
In this study, we explored the capabilities of scanning transmission electron microscopy tomography (STEM), nano X-ray computed tomography (nano-CT) and quantitative polarized Raman spectroscopy (qPRS) for the correlative structural and compositional analysis of bone. Freestanding bone micropillars (~25 µm diameter, ~55 µm height) were fabricated using femto-second laser ablation. The recently developed qPRS method (Kochetkova et al., 2020) allowed us to collect data on bone composition and MCF spatial orientation ~5 µm underneath the pillar surface with a spatial resolution of ~(0.3 µm)³. Laboratory phase contrast nano-CT tilt series were collected from selected volumes of interest up to 20 µm underneath the surface with an enhanced 3D resolution of ~(53 nm)³. Finally, sub-volumes were extracted by an established focused ion beam lift-out technique and imaged by STEM tomography. The quantitative local orientation of MCFs could be successfully estimated after 3D reconstruction and segmentation. The datasets were subsequently registered and correlated to each other to combine and cross validate the extracted information.
The combination of methods used in this study allows to resolve the ultrastructural organization of bone at the level of single MCF at a high resolution. The gained information may be used to identify novel biomarkers for bone quality, increasing the accuracy of bone fracture risk assessment in the future.