β-Galactosidase (β-Gal), as a glycoside hydrolase, is closely associated with cell senescence and primary ovarian cancer. However, there is still lack of facile and rapid sensing approach to monitor the β-Gal activity. In this work, a label-free and convenient sensing strategy to detect β-Gal activity has been proposed based on fluorescent graphene quantum dots (GQDs). In the presence of β-Gal, 4-nitrophenyl-β-D-galactopyranoside (NPGal) can be hydrolyzed into 4-nitrophenol (4-NP), which serves as a good quencher to quench the fluorescence of GQDs. The quenching mechanism is proven to be inner filter effect (IFE). Due to the specificity of the enzymatic reaction, this sensing method displays excellent selectivity and high sensitivity. A broad dynamic range from 20 to 200 U L and a detection limit of 4.4 U L for the β-Gal assay are achieved. Compared with the previously reported methods, this sensing strategy only needs one fluorescent nanomaterial without any modification and avoids time-consuming handling steps. Therefore, the sensing strategy based on fluorescent GQDs offers great potential for the recognition of disease-correlated enzyme activity.Copyright © 2020. Published by Elsevier B.V.