To determine biomechanical differences during single limb landing performed with and without shoe wear in anterior cruciate ligament reconstruction subjects.Barefoot and shod conditions were compared for both operated and contralateral limbs.Biomechanical analysis has been used to better understand the functional changes associated with anterior cruciate ligament reconstruction. However, no studies have investigated whether routine testing of patients should be performed barefoot or shod.A three-dimensional motion analysis and force plate system were used to obtain kinematic and kinetic data from eight anterior cruciate ligament reconstruction subjects during single-limb landings from a 15 cm vertical height.Peak knee flexion angles and moments were significantly reduced during barefoot testing in all eight subjects for the operated limb only (p < 0.01). The differences were however small; mean angular difference 3 degrees and mean moment difference 0.2 Nm/kg. Knee flexion angles at initial contact and peak vertical ground-reaction forces were not affected by shoe wear.These results show that shoe wear does affect some biomechanical variables on landing but not so as to affect comparisons between operated and contralateral limbs.Given the comparable results between conditions, biomechanical testing following anterior cruciate ligament reconstruction can be performed either barefoot or shod. Testing subjects barefoot has the advantage of easier marker placement, and alleviates the need for shoe wear standardization.