The effect of heat treatment on denaturation of Ara h1 protein, a major allergen from peanut, was studied using several techniques. Previously, Ara h1 protein was isolated from raw peanut using ammonium sulfate precipitation and chromatograpic techniques. Antibodies against Ara h1 protein were obtained in rabbits, conjugated with horseradish peroxidase, and used to develop a sandwich ELISA. Denaturation of Ara h1 protein was estimated by the loss of reactivity with its specific antibodies by ELISA. Kinetic and thermodynamic parameters of the denaturation process of Ara h1 protein were determined over a temperature range of 82-90 °C. Denaturation of Ara h1 was best described assuming a reaction order of 1.5. Thermal denaturation of Ara h1 protein was also studied by differential scanning calorimetry using several heating rates. The maximum peak temperature and the enthalpy of denaturation obtained by extrapolation to a scan rate of 0 °C/min were 90.22 °C and 1574 kJ/mol, respectively. The hydrophobicity of Ara h1 protein increased with the intensity of heat treatment, and aggregates were formed when the protein was treated at 90 °C for 10 min.