The invention discloses application of a molybdenum-based heteropolyacid-modified rare-earth fluorescent nanomaterial in glutathione assay. The method comprises the following steps: a series of glutathione solutions with normal concentrations respectively react with the molybdenum-based heteropolyacid-modified rare-earth fluorescent nanomaterial, so that a series of mixed liquors are obtained. Under the irradiation of near-infrared light, the changes of fluorescence intensities are detected; after a standard curve of fluorescence and glutathione concentrations is drawn, glutathione solution with an unknown concentration reacts with the molybdenum-based heteropolyacid-modified rare-earth fluorescent nanomaterial, so that a mixed liquor is obtained; under the irradiation of near-infrared light, the change of fluorescence intensity is detected and compared with the standard curve, and thereby the concentration of the glutathione solution is obtained. The method can quickly, sensitively and portably carry out quantitative assay on the glutathione, such as the quantitative assay of food, medicines, living samples and the like, moreover, the method is easy to operate, the linear effect is good, the limit of detection is low, the cost is low, instruments are convenient to carry, and movable assay can be implemented.