Background Trans-sacral axial L5/S1 interbody fusion (AxiaLIF), a novel surgical procedure, recently adopted in clinical practice, has excellent clinical outcomes. However, there is inadequate data on the feasibility of the approach in all adult patients and the optimal surgical approach is currently unclear; therefore, further studies are required. In order to enhance the surgical approach for AxiaLIF, prospective anatomical imaging optimization is necessary. The objective of this study was to investigate the ability of magnetic resonance imaging (MRI) to achieve an optimal procedural setting.Methods The subjects (n=40) underwent lumbosacral MRI examination. The median sagittal MRI images were analyzed and four measurement markers were defined as follows: the center of the L5/S1 disc (A), the anterior margin of the S1/2 disc space (B), the sacrococcygeal junction (C), and the coccygeal tip (D). The measurement markers were connected to each other to produce five lines (AB, AC, AD, BC, and BD), as reference lines for surgical approaches. The distance between each reference line and the anterior and posterior margins of the L5 and S1 vertebral bodies was measured to determine the safety of the respective approaches.Results In all patients, Lines AB and AC satisfied the imaging safety criteria. Line AB would result in a significant deviation from the median and was determined to be unsuitable for AxiaLIF. Line AD satisfied the imaging safety criteria in 39 patients. However, the anal proximity of the puncture point proved to be limiting. For lines BC and BD, the imaging safety criteria were satisfied in 70％ and 45％ of patients, respectively.Conclusions The AxiaLIF procedure is a safe technique for insertion of fusion implants in all subjects. Line AC is a favorable reference line for surgical approach and safe for all subjects, while line BC is not suitable for all subjects.