Abstract: Thermodynamic stability describes the tendency of protein unfolding, generally showed by the melting temperature (T_m) at which 50% of the protein is unfolded. In order to compare the detection characteristics and effects of different methods, differential scanning calorimetry (DSC), circular dichroism (CD) and differential scanning fluorimetry (DSF) were used to measure the T_m values of five proteins with different structural feature. The T_m values of different proteins gained using DSC, CD and DSF methods were compared. In summary, the T_m values of the same protein measured by the three methods were rather consistent, but differences were also observed. Additionally, proteins with complex structures did not show multiple T_m values.