Background We undertook a prospective clinical study to evaluate PCR.Ai’s (www.pcr.ai) accuracy and impact when automating the manual data-analysis and quality control steps associated with routine clinical pathogen testing using real-time PCR (qPCR). Objectives We evaluated the impact of PCR.Ai when used as the final interpretation/verification step for routine in-house qPCR tests for respiratory pathogens and for norovirus for a total of 22,200 interpretations. Study Design We compared PCR.Ai to our existing manual interpretation, to determine accuracy and hands-on time savings. PCR.Ai was accurate. Results and Conclusions There was 100% concurrence between validated respiratory virus and norovirus detection by our manual routine analysis method and PCR.Ai. Furthermore, there were significant routine savings with PCR.Ai of 45 min/respiratory run and 32 min/norovirus run. Our conclusion is that PCR.Ai is a highly accurate time-saving tool that reduces complexity of qPCR analysis and hence the need for specialists and hands-on time. It demonstrated capabilities to enable us to get results out more quickly with lower costs and less risk of errors.