Here we present extensive observations of stratospheric and upper tropospheric water vapor using the balloon-borne Cryogenic Frost point Hygrometer (CFH) in support of the Aura Microwave Limb Sounder (MLS) satellite instrument. Coincident measurements were used for the validation of MLS version 1.5 and for a limited validation of MLS version 2.2 water vapor. The sensitivity of MLS is on average 30% lower than that of CFH, which is fully compensated by a constant offset at stratospheric levels but only partially compensated at tropospheric levels, leading to an upper tropospheric dry bias. The sensitivity of MLS observations may be adjusted using the correlation parameters provided here. For version 1.5 stratospheric observations at pressures of 68 hPa and smaller MLS retrievals and CFH in situ observations agree on average to within 2.3% ± 11.8%. At 100 hPa the agreement is to within 6.4% ± 22% and at upper tropospheric pressures to within 23% ± 37%. In the tropical stratosphere during the boreal winter the agreement is not as good. The “tape recorder” amplitude in MLS observations depends on the vertical profile of water vapor mixing ratio and shows a significant interannual variation. The agreement between stratospheric observations by MLS version 2.2 and CFH is comparable to the agreement using MLS version 1.5. The variability in the difference between observations by MLS version 2.2 and CFH at tropospheric levels is significantly reduced, but a tropospheric dry bias and a reduced sensitivity remain in this version. In the validation data set a dry bias at 177.8 hPa of −24.1% ± 16.0% is statistically significant.