The aim of this study was the comparison of intraoperative kinematics, stability, and range of motion (ROM) between the native osteoarthritic knee and cruciate-substituting fixed-bearing total knee arthroplasty (TKA) using both an ultracongruent (UC) and a posterior-stabilized (PS) insert design in the same patient. We hypothesized less knee flexion and less antero-posterior stability in the UC TKA. Intraoperative measurements of kinematics, stability, and ROM were performed in 40 patients before soft-tissue release and bone cuts, and after implantation of the final femoral and tibial implants with both a UC insert and a PS insert. All measurements were performed using a navigation system. Kinematics changed significantly from a constant posterior femoral rollback before surgery to a paradoxical anterior translation during initial knee flexion with both inserts, but less pronounced with the PS insert (p < 0.001). There was significantly more posterior femoral rollback with the PS insert compared to the UC insert (p < 0.01). Stability measurements demonstrated no differences at full extension and 30° of knee flexion but significantly increased antero-posterior translation in 60° and 90° of knee flexion with the UC insert compared to the PS insert (p < 0.001). ROM measurements demonstrated improvement of knee flexion from 118° at the beginning of the surgery to 123° with the UC insert and 128° with the PS insert (p < 0.001). The use of a UC insert intraoperatively resulted in less antero-posterior stability and slightly less knee flexion compared to a PS insert. Surgeons should be aware of these differences when deciding for one of these options to substitute the posterior cruciate ligament (PCL). The impact of these findings on clinical outcome needs further investigation. II.