A small angle X-ray scattering (SAXS) synchrotron was used to evaluate 100 ovine m. longissimus lumborum, representing lamb (n = 50) and sheep (n = 50). The diffraction of X-rays gives information on muscle myofibril structure and fat content. The linear relationships between SAXS measures with measures such as, shear force,intramuscular fat content (IMF) and collagen content/solubility, were investigated. A relationship was found between the d-spacing of the actin/myosin fibril spacing (SAX1 and SAX2) and the cross sectional area of the rhombohedral unit cell (Cell area) and shear force after 1 and 5 day ageing. There was a positive relationship between IMF and a SAXS Fat area measure. There was a muscle site effect on SAX1, SAX2 and Cell area, with the cranial site having a larger distance between myofibrils. The potential of SAXS as a powerful research tool to determine not only the structural components of ovine tenderness, but also the fat content related to IMF is evident.