In this study, we carry out a theoretical investigation of the impact of transmission electron microscopy (TEM) sample thickness, which we varied from 2 to 140 nm, on the Si/SiO2 interface roughness that is directly extracted from a TEM image and is used for further calculation of the roughness scattering limited mobility (μsr) of Si metal-oxide-semiconductor (MOS) field-effect transistors [Zhao etal IEEE Trans. Electron Devices 57, 2055 (2010)]. We found that the values of the roughness height (Δ) and the correlation length (Λ) of the interface, as extracted from the TEM image, are underestimated, especially for the region in which the TEM sample thicknesses are large. Furthermore, we confirmed that the distribution function of the projected one-dimensional SiO2/Si interface is identical to that of the original two-dimensional interface. These results may help us to gain a more quantitative understanding of the roughness scattering limited mobility of advanced MOS devices by performing TEM measurements.