Pyrogens are components of microorganisms, like bacteria, viruses or fungi, which can induce a complex inflammatory response in the human body. Pyrogen contamination on medical devices prior operation is still critical and associated with severe complications for the patients. The aim of our study was to develop a reliable test, which allows detection of pyrogen contamination on the surface of medical devices. After in vitro pyrogen contamination of different medical devices and incubation in a rotation model, the human whole blood monocyte activation test (MAT), which is based on an IL-1β-specific ELISA, was employed. Our results show that when combining a modified MAT protocol and a dynamic incubation system, even smallest amounts of pyrogens can be directly detected on the surface of medical devices. Therefore, screening of medical devices prior clinical application using our novel assay, has the potential to significantly reduce complications associated with pyrogen-contaminated medical devices.