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The paper presents the investigation of thermal properties of thermal insulation chambers as an actuator in intelligent clothing, having the property of automatically adjusting the thermal protection level. The chambers are designed to vary their thickness based on the pressure of the inflated air in them. The pressure value measured in the thermal insulation chamber gives the microcomputer information on the thickness of the chamber. The paper presents the investigation of the functional dependencies of changes in the thickness of the chambers on the air pressure in them and the thermal resistance depending on the thickness of the thermal insulation chamber. Experimental thermal insulation chambers were made and integrated into an intelligent article of clothing and filled with air of 0–50 mbar, whereby chamber thicknesses of 0–25 mm were measured. Next, thermal resistance of 0.1876–0.5022 m2 k/W was measured on the thermal manikin. It was found that the ratio of thermal insulation of non-activated to maximally activated chambers was 1:2.7. Research has shown good results for the area where intelligent clothing can automatically adjust its thermal insulation properties. The technical systems described represent a suitable basis for experiments and scientific research during the introduction of intelligent clothing with active thermal protection into human life. The third-generation prototype shows very good properties from the aspect of automatic control of thermal protection in intelligent clothing. This forms the basis for further research. Cold protection has always been carried out by wearing garments with higher or lower thermal protection as well as wearing multiple layers of clothing. The conceptual starting point of the development of intelligent clothing is the development of an adaptive insulation layer with changeable thickness in the form of thermal insulation chambers filled with air. In this way, layered clothing as well as the use of several clothing layers can be avoided. Thus, one intelligent article of clothing regulating its thermal insulation properties can be used in a wide range of cold weather in the environment of the wearer.