Objective: Identification of parathyroid glands is often a challenge even for experienced surgeons. The feasibility and efficacy of fluorescence diagnosis for localization of parathyroids has already been proven in an experimental setting. In preparation for a clinical application of this technique in patients undergoing surgery for hyperparathyroidism, we evaluated the kinetics of fluorescence diagnosis. Design and methods: Fifty rats were randomized into eight groups with different photosensitization parameters using the photosensitizer aminolevulinic acid (ALA): a peritoneal lavage was performed with either 1.5 or 3.0% ALA solution and the induced photosensitization times varied from 0.5, 1 and 2 to 4 h. Under special fluorescence illumination, D-light, the exposed operative site with thyroid, parathyroid glands and neck muscles was examined. The identified parathyroid glands were studied according to fluorescence intensity by spectrometric measurement and compared with surrounding tissue.Results: Photosensitizer accumulation in parathyroid glands, indirectly measured by spectrometry, was up to 3.2 times higher than in thyroid and 2.6 times higher than in muscle tissue (2 h photosensitization with 3.0% ALA). Using 1.5% ALA, the optimum fluorescence intensity and ratio/contrast was slightly lower (parathyroid-to-thyroid ratio £ 3.0, parathyroid-to-muscle ratio £ 1.9) but was reached earlier (1 h) and hence considered as the parameter of choice for a clinical application. Conclusions: In future clinical application, intraoperative fluorescence diagnosis is expected to increase the ease of identification of atypically located or supernumeric glands. In combination with preoperative diagnostics, this may result in reduced operation time and avoidance of persistent hypercalcaemia.European Journal of Endocrinology 150 743-747