Land surface temperature (LST) is an important physical quantity at the land-atmosphere interface. Since 2016 the Collection 6 (C6) MODIS LST product is publicly available, which includes three refinements over bare soil surfaces compared to the Collection 5 (C5) MODIS LST product. To encourage the use of the C6 MODIS LST product in a wide range of applications, it is necessary to evaluate the accuracy of the C6 MODIS LST product. In this study, we validated the C6 MODIS LST product using temperature-based method over various land cover types, including grasslands, croplands, cropland/natural vegetation mosaic, open shrublands, woody savannas, and barren/sparsely vegetated. In situ measurements were collected from various sites under different atmospheric and surface conditions, including seven SURFRAD sites (BND, TBL, DRA, FPK, GCM, PSU, and SXF) in the United States, three KIT sites (EVO, KAL, and GBB) in Portugal and Namibia, and three HiWATER sites (GBZ, HZZ, and HMZ) in China. The spatial representativeness of the in situ measurements at each site was separately evaluated during daytime and nighttime using all available clear-sky ASTER LST products at 90m spatial resolution. Only six sites during daytime are selected as sufficiently homogeneous sites despite the usually high spatial thermal heterogeneity, whereas during nighttime most sites can be considered to be thermally homogeneous and have similar LST and air temperature. The C6 MODIS LST product was validated using in situ measurements from the selected homogeneous sites during daytime and nighttime: except for the GBB site, large RMSE values (> 2 K) were obtained during daytime. However, if only satellite LST with a high spatial thermal homogeneity on the MODIS pixel scale are used for LST validation, the best daytime accuracy (RMSE<1.3 K) for the C6 MODIS LST product is achieved over the BND and DRA sites. Except for the DRA site, the RMSE values during nighttime are<2 K at the selected homogeneous sites. Furthermore, the accuracy of the C6 MODIS LST product was compared with that of the C5 MODIS LST product during nighttime at the selected homogeneous sites. Except for the GBB site, there are only small differences (< 0.4 K) between the RMSE values for the C5 and C6 MODIS LST products.