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Abstract β -Galactosidase (β -gal) is an important biomarker for primary ovarian cancers and cell senescence; however, a fast response fluorescent probe for ratiometric monitoring is still rare. A novel, ratiometric water-soluble fluorescent probe (FLM) for β -gal was developed. The emission ratio F 550 / F 450 reached the maxima at about 5 min and can be used for real-time detection of β -gal; the ratio gained an ultimate enhancement of about 260-fold. The ratio (F 550 / F 450) displayed brilliant β -gal-dependent performance and responded linearly with β -gal activity. The probe showed wonderful biocompatibility and was successfully used for the bioimaging of endogenous β -gal in the human ovarian cancer cell line OVCAR-3. Graphical abstract fx1 Highlights • A novel ratiometric fluorescent probe for β -galactosidase was developed. • The probe can respond quickly to β -galactosidase in PBS buffer. • The fluorophore FL has a high fluorescence quantum yield in PBS buffer. • The probe possesses excellent biocompatibility and low cytotoxicity. • The probe can be used for real-time visualization of β -galactosidase in living cells. [ABSTRACT FROM AUTHOR]