Objective: To validate an amended protocol for clinical diagnosis of sporadic Creutzfeldt-Jakob Disease (sCJD) including Real-Time Quaking-induced Conversion (RT-QuIC) and to observe its utilization in CJD surveillance. Methods: In the framework of a prospective epidemiological study, all neuropathological confirmed sCJD cases that received CSF RT-QuIC analysis during diagnostic work up (n=65) and a control group of non- CJD cases (n=118) were selected to investigate the accuracy of an amended diagnostic protocol. The patients had been referred to the German National Reference Center for Transmissible Spongiform Encephalopathies. The influence of the amended protocol on incidence figures was evaluated in the context of three years of surveillance activity (screened cases using 14-3-3 test: n=18.789, highly suspicious cases of CJD: n=704). Annual incidences were calculated using current criteria and the amended protocol. Results: The amended protocol showed a sensitivity of 97% and a specificity of 99%. When it was applied to all suspected cases that were referred to the reference center, the assessed incidence of CJD increased from 1.7 to 2.2 per million in 2016. Conclusion: CJD surveillance remains challenging as information from external healthcare institutions can be limited. RT-QuIC shows excellent diagnostic accuracy when applied in the clinical setting to symptomatic patients. Data for RT-QuIC alone, when applied as a general screening test, are not available yet. We propose an amended research protocol which improves early and accurate clinical diagnosis of sCJD during surveillance activities. The utilization of this protocol will probably lead to a significant increase of the incidence rate.