Silica glass is frequently used as a device material for micro/nano fluidic devices due to its excellent properties, such as transparency and chemical resistance. Wet etching by hydrofluoric acid and dry etching by neutral loop discharge (NLD) plasma etching are currently used to micromachine glass to form micro/nano fluidic channels. Electro-osmotic flow (EOF) is one of the most effective methods to drive liquids into the channels. EOF mobility is affected by a property of the micromachined glass surfaces, which includes surface roughness that is determined by the manufacturing processes. In this paper, we investigate the effect of micromaching processes on the glass surface topography and the EOF mobility. We prepared glass surfaces by either wet etching or by NLD plasma etching, investigated the surface topography using atomic force microscopy, and attempted to correlate it with EOF generated in the micro-channels of the machined glass. Experiments revealed that the EOF mobility strongly depends on the surface roughness, and therefore upon the fabrication process used. A particularly strong dependency was observed when the surface roughness was on the order of the electric double layer thickness or below. We believe that the correlation described in this paper can be of great help in the design of micro/nano fluidic devices.