In this research, the thermal comfort properties of a newly known fiber called Estabragh (Milkweed) have been investigated. Estabragh is a natural hollow fiber which is expected to provide excellent insulation properties due to their individual characteristics. Thermal comfort is one of the important fabric properties especially in the area of sport application which is considered in this research. For investigating this, 50–50% Estabragh/cotton-blended spun yarns was produced using short-staple spinning system. Two different sets of interlock-knitted samples using 100% cotton and Estabragh/cotton-blended yarns were produced individually on a flat knitting machine. Heat transfer rate, water vapor transmission, air permeability, and capillary absorption of produced fabrics were measured and compared. The results revealed that the vapor transmission rate through the knitted samples produced from Estabragh/cotton-blended yarns was 66.4% which is higher than that of the 100% cotton samples as 60.22%. It was also found that 100% cotton samples exhibited less moisture transfer rate than cotton/Estabragh samples. As far as heat transfer is concerned, it was concluded that the heat transfer ability of 100% cotton samples is higher than that of the Estabragh/cotton samples by about 47%. It was also shown that the capillary absorption of samples produced from Estabragh/cotton-blended yarns was higher than that of the 100% cotton samples. Compared to the 100% cotton samples, air permeability of Estabragh/cotton-knitted structures was found to be relatively low. The results totally proved that Estabragh fibers have a lot of advantages in order to be used in different fields of applications especially in sport areas.