Interaction of clenbuterol hydrochloride(CL) with human immunoglobulin G(HIgG) was studied using fluorescence quenching spectroscopy，three-dimensional fluorescence spectroscopy，UV-vis absorption spectroscopy and circular dichroism(CD) spectroscopy under simulative physiological conditions for the first time.The binding parameters and the thermodynamic parameters for the reaction of CL with HIgG at different temperatures were calculated according to Scatchard equation and Van-t Hoff equation，respectively.Experimental results of the fluorescence quenching spectra showed that the fluorescence intensity of HIgG was quenched by the gradual addition of CL.The binding constants of CL with HIgG at 298，304,310 K were calculated to be 2.95×104，2.35×104,1.82×104 L/mol，respectively，meaning that the quenching mechanism was a static quenching.Meanwhile，the corresponding numbers of binding sites(n) were calculated to be 0.84，0.87 and 0.94，respectively.The thermodynamic parameters of the reaction，namely standard enthalpy ΔH0 and entropy ΔS0，were calculated to be-30.47 kJ?mol-1 and-16.58 J?K?mol-1，respectively，suggesting that there were several forces in the binding of CL with HIgG,and the hydrogen bond and Vander Waals force were the predominant intermolecular forces in stabilizing the CL-HIgG complex.Experimental results obtained from the three-dimensional fluorescence spectroscopy and UV-vis absorption spectroscopy confirmed that the secondary structure of HIgG was altered in the presence of CL in aqueous solution.Furthermore，a decrease of the β-structure compositions of HIgG was observed by CD spectroscopy when the molar concentration ratio of HIgG to CL was 1∶4.