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Description: [This project is now a published open access article in Scientific Reports: https://www.nature.com/articles/s41598-021-00731-7] In electroencephalography (EEG), microstates are distributions of activity across the scalp that persist for several tens of milliseconds before changing into a different topographical pattern. Microstate analysis is a promising way of utilizing EEG as both temporal and spatial imaging tool, but has mostly been applied to resting state data. This study aimed to conceptually replicate microstate findings of valence and arousal processing and to investigate the effects of emotion regulation on microstates, using existing data of an EEG paradigm with 107 healthy adults who were to actively view emotional pictures, cognitively detach from them, or suppress facial reactions. EEG data were clustered into microstates based on topographical similarity and compared on global and electrode level between conditions of interest. Within the first 600 ms after stimulus onset only the comparison of viewing positive and negative pictures yielded significant global results, caused by different electrodes depending on the microstate. Since the microstates associated with more and less arousing pictures did not differ from each other, sequential processing of valence and arousal information could not be replicated. When extending the analysis to 2,000 ms after stimulus onset, global microstate differences were exclusive to the comparison of viewing and detaching from negative pictures. Intriguingly, we observed the novel phenomenon of a significant global difference that could not be attributed to single electrodes on the local level. This suggests that microstate analysis can detect differences beyond those detected by event-related potential analysis, simply by not confining the analysis to a few electrodes.