Anterior cruciate ligament (ACL) reconstruction is a significant clinical problem. Clinical studies reveal that 11%–32% of patients show an unsatisfactory prognosis and that up to 10% may require surgical revision. Graft-tunnel healing is one of the major factors affecting the outcome of ACL reconstruction. Given that there is no analog like tendon insertion in the bone tunnel in animal or human, it is necessary to establish an appropriate animal model for a better understanding of the biology of graft-tunnel healing; in particular, the structure of the tendon insertion site in the bone tunnel should be known in prior. Recently, a protocol for establishing ACL reconstruction animal models has been developed after a critical review of the literature in the past decades. The assessment protocol consists of three-dimensional (3D) structural analysis of bone ingrowth by micro-computed tomography (micro-CT), two-dimensional (2D) structural analysis of newly formed tendon insertion in the bone tunnel by routine histology, and mechanical testing of the strength of the graft–tunnel complex as the endpoint evaluation. Densitometric evaluation is also used to assess the changes of pre-existing bone with graft–tunnel healing by peripheral quantitative computed tomography (pQCT). In this chapter, we also discuss several aspects on the application of this experimental model for developing therapeutic strategies to enhance graft–tunnel healing.