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A better knowledge of the kinematics behavior of total knee replacement (TKR) during activity still remains a crucial issue to validate innovative prosthesis designs and different surgical strategies. Tools for more accurate measurement of in vivo kinematics of knee prosthesis components are therefore fundamental to improve the clinical outcome of knee replacement. In the present study, a novel model-based method for the estimation of the three-dimensional (3-D) position and orientation (pose) of both the femoral and tibial knee prosthesis components during activity is presented. The knowledge of the 3-D geometry of the components and a single plane projection view in a fluoroscopic image are sufficient to reconstruct the absolute and relative pose of the components in space. The technique is based on the best alignment of the component designs with the corresponding projection on the image plane. The image generation process is modeled and an iterative procedure localizes the spatial pose of the object by minimizing the Euclidean distance of the projection rays from the object surface. Computer simulation and static/dynamic in vitro tests using real knee prosthesis show that the accuracy with which relative orientation and position of the components can be estimated is better than 1.5° and 1.5 mm, respectively. In vivo tests demonstrate that the method is well suited for kinematics analysis on TKR patients and that good quality images can be obtained with a carefully positioning of the fluoroscope and an appropriate dosage. With respect to previously adopted template matching techniques, the present method overcomes the complete segmentation of the components on the projected image and also features the simultaneous evaluation of all the six degrees of freedom (DOF) of the object. The expected small difference between successive poses in in vivo sequences strongly reduces the frequency of false poses and both the operator and computation time.