The characteristics of host-guest complexation between cucurbit ［7］ uril (CB7) and methylene blue (MB) were investigated by fluorescence spectrometry in acetate buffer solution at room temperature. It was found that the fluorescence intensity of methylene blue regularly increased upon the addition of cucurbit ［7］ uril accompanying with a blue-shift of the position of the emission maximum. The results indicate the formation of complex between CB7 and MB at a 1∶1 complex stoichiometry and the association constant was calculated by applying a deduced equation. 1H NMR spectra were applied to to complement the fluorescence work to verify the formation of the complex. From the temperature dependence of the equilibrium constants, thermodynamics parametersH andS values were obtained, indicating an enthalpic driving force for complexation. The possible interaction mechanism was also discussed. This work may extend the application range of cucurbit ［7］ uril in biochemistry and pharmaceutical analysis.