Albumin Binding Enhances the Lysosomal β-Galactosidase Sensitivity of Fluorogenic Probes Characteristic of Intramolecular Charge Transfer

Glycosidases generally function in specific organelles to hydrolyze glycoconjugates. Thus, the in situ visualization of glycosidase activities in an organelle-targeted manner can help to better delineate their biological functions. Lysosomal β-galactosidase (β-Gal) is reported to be a biomarker for ovarian cancer and cellular senescence. Here, we developed near-infrared fluorogenic probes for the detection of β-Gal activity based on modulating the intramolecular charge transfer (ICT) of a hemicyanine dye. The constructed probe, Hcy-Lyso-Gal bearing a morpholine group to target the lysosomes, enabled the visualization of lysosomal β-Gal activity in different live cells. Through a series of experiments, we rationalized that human serum albumin binding could be the main reason by which to significantly enhance the fluorescence of the probe in the acidic lysosomes where its phenol anion is protonated to quench dye fluorescence. In addition, the probe was used to image cell lines with different endogenous β-Gal expression levels and visualize lysosomal β-Gal in senescent cells. This study offers insight into the employment of ICT-based probes for fluorescence-based imaging of lysosomal enzymes.