The purpose of this study was to evaluate the usefulness of 99mTc labelled ciprofloxacin imaging in detecting the presence of infection in patients with symptomatic knee prostheses. Among 16 randomly selected patients of whom seven had infection based on clinical and microbiological findings and nine did not, 99mTc-ciprofloxacin images were obtained at 1, 4 and 24h after the injection of the tracer. While there was some diffuse non-specific accumulation of 99mTc-ciprofloxacin in large synovial joints and in prosthetic knee joints, the infected knee prostheses were found to show more intensive focal uptake, which also extended outside the synovial cavity. The infection related uptake remained visible in the 24h images, whereas non-specific uptake had a fading tendency at this time point. 99mTc-ciprofloxacin imaging showed diagnostic sensitivity of 86% and a specificity of 78% for correctly classifying the presence of infection. The data indicate that 99mTc-ciprofloxacin imaging may be used in the diagnosis of knee prosthesis infections. Infection-related uptake remains visible in the 24h images and is typically found also outside the synovial cavity, which should be noted in the evaluation of the images.