Isothermal chamber is filled with a certain density of high thermal conductivity porous material, which has a wide range of applications in the flow measurement of pneumatic field, and its isothermal characteristic is critical to its application results. In this paper, in order to improve the numerical calculation accuracy of the isothermal characteristics during discharge process, a discharge model of isothermal chamber with fractal effective thermal conductivity (ETC) for determining the isothermal characteristic is reported. Firstly, the stuffer in isothermal chamber is considered as porous random fiber bundle, and an ETC prediction model of anisotropic porous random fiber bundle in both vertical and horizontal directions is established by fractal theory. This model with two directions is in good agreement with the experimental results, and the relative root mean square errors (RRMSE) are 3.94% and 9.85%, respectively. Secondly, the discharge model with fractal ETCs is built, and the isothermal characteristics of isothermal chambers with three different porosities are determined by numerical simulation. Finally, experiments to determine the isothermal characteristic are carried out. The numerical simulation results are in good agreement with the experimental results, and the relative errors are less than 3%. It could be concluded that accurately determining the ETC of the stuffer in isothermal chamber can improve the numerical calculation accuracy of the isothermal characteristic. Moreover, compared with the experimental method, numerical method is energy-saving and timesaving.