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A self-immobilizing fluorescent probe for single-cell resolution imaging of senescence-associated β-galactosidase
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DOI：10.1016/j.dyepig.2026.113734.png)
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Abstract
En 中文
Senescence-associated beta-galactosidase (SA-beta-gal) is an endogenous lysosomal enzyme overexpressed in senescent cells and thus widely used as a significant biomarker of cellular and tissue senescence. However, the application of existing fluorescent probes for SA-beta-gal detection is often hampered by their tendency to leak out of senescent cells after enzymatic hydrolysis. Herein, we develop a self-immobilizing SA-(3-gal probe TOF-beta gal, which generates a quinone methide intermediate after enzymatic cleavage that covalently binds to nearby proteins through bioorthogonal reactions, thereby enabling in situ detection of SA-beta-gal at single-cell resolution. TOF-beta gal integrates an environment-sensitive thiazole orange fluorophore, a beta-galactosidase-recognizing glycosidic moiety, and an ortho-fluoromethyl group as a protein-anchoring precursor. The probe displays minimal background fluorescence in aqueous media but exhibits a significant turn-on response in the simultaneous presence of A. oryzae (3-gal and bovine serum albumin (BSA). Furthermore, it shows excellent selectivity and a quantitative response to A. oryzae beta-gal in vitro, with a linear detection range from 0.1 to 1 U/mL and a detection limit as low as 0.012 U/mL. Moreover, TOF-beta gal is capable of monitoring endogenous SA-beta-gal activity in living cells, distinguishing senescent cells from non-senescent cells and quantifying the degree of senescence. These results unambiguously demonstrate that TOF-beta gal is a promising tool for the imaging of senescence.
Keywords:
Fluorescent probes
Senescence-associated (3-galactosidase
Single-cell resolution
Imaging
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4.2
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2.9W
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