Rotational kinematics of the knee is not fully restored after single-bundle anterior cruciate ligament (ACL) reconstruction. Cadaveric experiments using knee testing machines have suggested anatomical reconstruction replacing the anteromedial and posterolateral bundles could restore knee kinematics more effectively than single-bundle reconstruction. However, practical tools to objectively assess knee rotational laxities clinically have not been available. We used an optically based computer-assisted navigation system to measure the tibiofemoral motion kinematics in four fresh whole cadavers. Standard clinical knee laxity tests (anterior drawer, Lachman, and pivot shift) were performed and the kinematics described in terms of tibial axial rotation and anteroposterior translation. Data were obtained for intact knees after excision of the ACL and sequential reconstruction of the anteromedial and posterolateral bundles. In the ACL-deficient knee, the mean maximum tibial rotation during the pivot shift test was 27° and mean maximum translation 11 mm. Reconstruction of the anteromedial bundle reduced the rotational component to 18° and translation to 7 mm. Reconstruction of the posterolateral bundle reduced rotation to 14°. This pilot study suggests computer assisted navigation could provide a practical method to objectively measure the pivot shift and may be used clinically to demonstrate differences in the control of tibiofemoral rotation kinematics afforded by single and two-bundle ACL reconstructions.