Determination of ethanol-gasoline blends is of practical importance, as a high content of ethanol will significantly accelerate the corrosion of metallic materials in gasoline vehicle engines. However, the existing fluorescence detection method should draw support from excitation-emission matrix fluorescence technique combined with chemometric algorithms for data analysis. In the work, we report a novel fluorescent method by applying an environment-sensitive fluorescent dye for determination of ethanol-gasoline blends from a single emission spectrum, making fluorescence detection much more user-friendly.