A turn on fluorescent assay for real time determination of β-galactosidase and its application in living cell imaging

Abstract

In recent years, fluorescent probes based on chemical reactions have been widely investigated as a powerful and noninvasive method for the diagnosis of diseases. β-Galactosidase (β-gal), a typical lysosomal glycosidase, over expressed in senescent cells and primary ovarian cancer cells, which has been considered as an important biomarker cell senescence and primary ovarian cancers. Fluorescent probes for the determination of β-gal provide an excellent choice for visualization of cell senescence. In this work, a turn on fluorescent probe (HBT-gal) for β-gal activity was developed based on the enzymatic hydrolysis of glycosidic bonds. HBT-gal showed little fluorescence in aqueous buffer excited at 415 nm, while emitted green fluorescence centered at ∼ 492 nm upon incubated with β-gal. The sensing scheme showed high selectivity and sensitivity for β-gal activity with a limit of detection calculated as low as 0.19 mU/mL. Moreover, HBT-gal was successfully applied to image β-gal activity in senescent Hep G2 cells treated with H₂O₂. Therefore, probe HBT-gal demonstrated a potential usage for the determination of cell senescence using β-gal as a biomarker.

Keywords: Biological imaging; Fluorescence receptor; Real time detection; Senescent cells; β-galactosidase.