In this study, the influence of illumination disturbances on surface topography measurements performed using Focus Variation (FV) microscopy was investigated. Experiments were carried out on a sinusoidal roughness standard measured with an Alicona μCMM system under varying illumination conditions. The results show that seemingly negligible ceiling illumination significantly distorts the 2D microscopic image and markedly degrades the quality of the reconstructed 3D topography. Although the FV algorithms largely preserve the global directional character of the surface, the data exhibit increased noise and a pronounced rise in the number of non-measured points (NM), exceeding a tenfold increase compared to the reference conditions. The Ssk/Rsk and Sp/Rp parameters were found to be particularly sensitive to illumination disturbances, with Sp/Rp deviating by more than 46% under ceiling illumination. The combined illumination configuration mitigates some of these effects, improving the quality of the 3D data due to the dominant contribution of coaxial light. In summary, the study highlights that seemingly insignificant ambient light sources may constitute a significant and often overlooked source of measurement error in FV-based surface metrology.