A practical guide for in situ mechanical testing of musculoskeletal tissues using synchrotron tomography

Musculoskeletal tissues are complex hierarchical materials where mechanical response is linked to structural and material properties at different dimensional levels. Therefore, high-resolution three-dimensional tomography is very useful for assessing tissue properties at different scales. In particular, Synchrotron Radiation micro-Computed Tomography (SR-microCT) has been used in several applications to analyze the structure of bone and biomaterials. In the past decade the development of digital volume correlation (DVC) algorithms applied to SR-microCT images and its combination with in situ mechanical testing (four-dimensional imaging) have allowed researchers to visualise, for the first time, the deformation of musculoskeletal tissues and their interaction with biomaterials under different loading scenarios. However, there are several experimental challenges that make these measurements difficult and at high risk of failure. Challenges relate to sample preparation, imaging parameters, loading setup, accumulated tissue damage for multiple tomographic acquisitions, reconstruction methods and data processing. Considering that access to SR-microCT facilities is usually associated with bidding processes and long waiting times, the failure of these experiments could notably slow down the advancement of this research area and reduce its impact. Many of the experimental failures can be avoided with increased experience in performing the tests and better guidelines for preparation and execution of these complex experiments; publication of negative results could help interested researchers to avoid recurring mistakes. Therefore, the goal of this article is to highlight the potential and pitfalls in the design and execution of in situ SR-microCT experiments, involving multiple scans, of musculoskeletal tissues for the assessment of their structural and/or mechanical properties. The advice and guidelines that follow should improve the success rate of this type of experiment, allowing the community to reach higher impact more efficiently.