Background: Anterior cruciate ligament (ACL) graft conditioning protocols to decrease postoperative increases in anterior tibial translation and pivot-shift instability have not been established. Purpose: To determine what ACL graft conditioning protocols should be performed at surgery to decrease postoperative graft elongation after ACL reconstruction. Study Design: Controlled laboratory study. Methods: A 6 degrees of freedom robotic simulator evaluated 3 ACL graft constructs in 7 cadaver knees for a total of 19 graft specimens. Knees were tested before and after ACL sectioning and after ACL graft conditioning protocols before reconstruction. The ACL grafts consisted of a 6-strand semitendinosus-gracilis TightRope, bone–patellar tendon–bone TightRope, and bone–patellar tendon–bone with interference screws. Two graft conditioning protocols were used: (1) graft board tensioning (20 minutes, 80 N) and (2) cyclic conditioning (5°-120° of flexion, 90-N anterior tibial load) after graft reconstruction to determine the number of cycles needed to obtain a steady state with no graft elongation. After conditioning, the grafts were cycled a second time under anterior-posterior loading (100 N, 25° of flexion) and under pivot-shift loading (100 N anterior, 5-N·m internal rotation, 7 N·m valgus) to verify that the ACL flexion-extension conditioning protocol was effective. Results: Graft board tensioning did not produce a steady-state graft. Major increases in anterior tibial translation occurred in the flexion-extension graft-loading protocol at 25° of flexion (mean ± SD: semitendinosus-gracilis TightRope, 3.4 ± 1.1 mm; bone–patellar tendon–bone TightRope, 3.2 ± 1.0 mm; bone–patellar tendon–bone with interference screws, 2.4 ± 1.5 mm). The second method of graft conditioning (40 cycles, 5°-120° of flexion, 90-N anterior load) produced a stable conditioned state for all grafts, as the anterior translations of the anterior-posterior and pivot-shift cycles were statistically equivalent ( P < .05, 1-20 cycles). Conclusion: ACL graft board conditioning protocols are not effective, leading to deleterious ACL graft elongations after reconstruction. A secondary ACL graft conditioning protocol of 40 flexion-extension cycles under 90-N graft loading was required for a well-conditioned graft, preventing further elongation and restoring normal anterior-posterior and pivot-shift translations. Clinical Relevance: There is a combined need for graft board tensioning and robust cyclic ACL graft loading before final graft fixation to restore knee stability.