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Even though atmospheric-pressure plasma jet (APPJ) has been shown highly effective in many biomedical applications because of reactive oxygen/nitrogen species (RONS) generated in the discharge, free jet region, and at the interfacial region with the treated surface. However, most of the studies characterized APPJ only in either discharge or free jet region. In this paper, we measured optical emissions of RONS in free jet and plasma/surface interfacial regions of a helium-based dielectric barrier discharge APPJ. The results show that the emissions caused by RONS (nitric oxide, hydroxyl radical, and atomic oxygen) are enhanced 3–5 times at plasma/surface interfacial region as compared with those of free jet region. Some typical sterilization experiments using the current helium-based APPJ show that this helium-based APPJ effectively kills the oral bacteria (S. mutans) in 15 s, which was caused by the cell wall disruption. We believe that measuring the properties at plasma/surface interfacial region is more important than in the free jet region in characterizing APPJ for most realistic applications. [ABSTRACT FROM AUTHOR]