Thermal and moisture behavior is crucial in determining the comfort of clothing. Surface modification of fibers brings new functions and properties to fibrous materials, but the comfort of the as-made clothing changes accordingly due to different thermal and moisture behaviors. In this study, physical and chemical modifications have been conducted to wool fabric to investigate their effects on the thermal and moisture behaviors of clothing. Surface wettability, wicking properties, thermal and moisture resistance together with surface temperature and humidity of the microclimate have been tested. It has been found that the wettability of wool fabric is greatly improved and wicking properties are granted to the fabric. Higher thermal and moisture resistance have been noticed for the treated wool fabric. The treated fabric shows longer moisture liberation process and the surface temperature and humidity varies in a different way compared with untreated fabric. Understanding the thermal and moisture behaviors of fabric after surface modification would benefit the further development of surface functionalizing technology and design of next to skin clothing.