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Radiographs of 20 plastinated human cadaveric lower legs were obtained in 12 positions of rotation to determine the optimal parameter for reliable assessment of syndesmotic and ankle integrity, and to assess the effect of positioning of the ankle on this parameter. Three observers measured eight parameters twice after four repetitions of ankle positioning. Intraclass correlation coefficients and reproducibility were assessed. Some tibiofibular overlap was present in all radiographs in any position of rotation. The medial clear space was smaller than or equal to the superior clear space in all radiographs. Intraclass correlation coefficients of the other parameters were too weak for reliable quantitative measurements, as was shown with a mixed model analysis of variance. This resulted from the inability to reproduce ankle positioning, even under optimal laboratory circumstances. This study shows that no optimal radiographic parameter exists to assess syndesmotic integrity. Tibiofibular overlap and medial and superior clear space are the most useful, because one-sided traumatic absence of tibiofibular overlap may be an indication of syndesmotic injury, and a medial clear space larger than a superior clear space is indicative of deltoid injury. Additional quantitative measurement of all syndesmotic parameters with repeated radiographs of the ankle cannot be done reliably and therefore are of little value.