Effect of soft tissue sample preparation techniques for scanning small-angle X-ray scattering experiments.

Scanning small-angle X-ray scattering (sSAXS) has found multiple applications as a technique to probe the nanostructure in soft tissues and pathologies thereof. However, fresh tissue is fragile and prone to the quick onset of decomposition and autolysis. It lacks the firmness required for uniform and thin sectioning, resulting in the loss of 2D resolution offered by focused X-ray beams, because the signal would be integrated through thick and/or irregular sections. Tissue processing, that includes fixation and embedding, is used to mitigate these issues but can by itself introduce structural changes in the tissues and impede the correct interpretation of sSAXS data. Here the extent of these structural changes in the SAXS signal caused by common tissue preservation methods on the example of skeletal muscle tissue, consisting of both muscle and surrounding connective tissue, was studied. This can guide an informed choice of preservation method tailored for specific experimental requirements. While some techniques performed better than others, all tissue-processing methods induced structural changes to a certain degree. The choice of preservation method is therefore a balance between sectioning requirements and type of tissue used, as well as targeted structural information.