Abstract Wafer bonding is an essential technology for attaching various materials, including glass. Glass-glass bonding has been applied to fabricate glass micro/nanofluidic devices that work as analytical tools for chemical and biological research. Here, we report the fabrication of micro/nanofluidic devices consisting of a high refractive index (HRI) glass as a component of the device for the first time. Although HRI glass has been used for high-sensitivity and high-resolution imaging in microscopy, it has not been used for glass fluidic devices due to the lack of bonding method for HRI glass. The glass fluidic devices were made through glass-glass bonding at room temperature, which could avoid severe damage to bonded substrates of two glass types due to different rates of thermal expansion. The pressure endurance of these devices was measured to find the correlation with the surface properties of glass substrates. The measured results indicated that chemical groups other than SiOH enhanced the pressure endurance of the devices. Thus, these chemical groups helped to form the fluidic devices consisting of HRI glass and another glass type, borosilicate or fused silica, to achieve practical pressure endurance up to 270 kPa. The HRI glass-incorporating device showed 8-fold higher sensitivity and 30 % narrower spatial resolution of fluorescence signals than the other devices with non-HRI glass.