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Background: A comparison of natural versus replaced tibio-femoral kinematics in vivo during challenging activities of daily living can help provide a detailed understanding of the mechanisms leading to unsatisfactory results and lay the foundations for personalised implant selection and surgical implantation, but also enhance further development of implant designs towards restoring physiological knee function. The aim of this study was to directly compare in vivo tibio-femoral kinematics in natural versus replaced knees throughout complete cycles of different gait activities using dynamic videofluoroscopy., Methods: Twenty-seven healthy and 30 total knee replacement subjects (GMK Sphere, GMK PS, GMK UC) were assessed during multiple complete gait cycles of level walking, downhill walking, and stair descent using dynamic videofluoroscopy. Following 2D/3D registration, tibio-femoral rotations, condylar antero-posterior translations, and the location of the centre of rotation were compared., Findings: The total knee replacement groups predominantly experienced reduced tibial internal/external rotation and altered medial and lateral condylar antero-posterior translations compared to natural knees. An average medial centre of rotation was found for the natural and GMK sphere groups in all three activities, whereas the GMK PS and UC groups experienced a more central to lateral centre of rotation., Interpretation: Each total knee replacement design exhibited characteristic motion patterns, with the GMK Sphere most closely replicating the medial centre of rotation found for natural knees. Despite substantial similarities between the subject groups, none of the implant geometries was able to replicate all aspects of natural tibio-femoral kinematics, indicating that different implant geometries might best address individual functional needs., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)