The influence of environmental factors on dimensional measurements of atomic force microscopy (AFM) was investigated experimentally. Measurements were taken with environmental control over a whole AFM chamber and a local sample chamber to highlight the influence of working conditions on the instrument itself. Both temperature and humidity were found to have a significant impact on pitch measurements of a two-dimensional grating. The effect of temperature on the behavior of the microscope itself is generally larger than the thermal expansion or contraction of the sample. The effect of humidity was further determined to be relevant to the scan direction and velocity. For precise AFM dimensional measurements, the possible influences of temperature and humidity must be carefully considered.