Purpose: The purpose of this study was to evaluate the isometry of an anatomic femoral tunnel and anterior tibial tunnel positions. Methods: Tibial tunnels were made at 2 different locations in 10 cadaveric knees: the conventional tunnel and a more anterior position. Three-dimensional computed tomography (CT) scanning was then performed at 0 degrees, 30 degrees, 60 degrees, 90 degrees, and 120 degrees. After removal of the anterior cruciate ligament from its femoral attachment, the 2 different femoral tunnels were marked at (1) the vertical femoral tunnel point and (2) the anatomic femoral tunnel point. After scans were repeated for coordinate transformation, the change in length between the tunnels was calculated with imaging software (OsiriX, version 3.2; Apple, Cupertino, CA) and the center of rotation for the femoral tunnels was calculated with a least squares fitting algorithm. Results: The conventional tibial tunnel-vertical femoral tunnel combination showed the least excursion as knee flexion angle changed. The vertical femoral tunnel combination groups showed a trend toward increasing length as the knee flexion angle increased. In contrast, the anatomic femoral tunnel combination groups displayed a trend toward decreased length with increasing knee flexion. At less than 30 degrees of flexion, the tibial anterior-anatomic femoral tunnel showed the least excursion. Conclusions: The anatomic femoral tunnel was nonisometric, and the differences in isometry for each tunnel type were explained primarily by differences in relations between the centers of rotation of tunnels and tunnel position. When a femoral anatomic tunnel is chosen for anterior cruciate ligament reconstruction, the anterior tibial tunnel offers greater isometric benefits than the conventional tibial tunnel, especially in near full extension. Clinical Relevance: The distance between anatomic femoral and tibial tunnels is greatest in full extension and decreases with flexion. This would result in graft laxity. The surgeon should give consideration to a more anterior tibial tunnel position, which shows less excursion in early flexion.