Dual-Channel Phosphorescence Ratiometry and Phosphorescence Lifetime Imaging of Mitochondria-Specific Methionine Sulfoxide Reductase Activity

Abstract

Methionine sulfoxide reductases (Msrs) are essential for preserving redox homeostasis in the nervous system, with dysregulation implicated in Alzheimer’s disease (AD). Conventional fluorescence-based assays for Msrs activity sensing are hampered by background interference, limited sensitivity, and inadequate quantification. This work introduces a novel supramolecular probe exhibiting redox-responsive dual-channel room-temperature phosphorescence (RTP) in aqueous media on a microsecond time scale. Upon reduction by Msrs, the probe transitions from its oxidized to reduced state, manifested by a red-shifted phosphorescence emission and extended lifetime in the microsecond range, which enables precise quantification of mitochondria-targeted Msrs activity via phosphorescence ratiometry and phosphorescence lifetime imaging (PLIM). The probe’s utility is demonstrated in visualizing neuronal Msrs activity and distribution within the mouse brain, which reveals a marked downregulation of Msrs activity in an AD model, highlighting the probe’s potential in elucidating redox-related pathological mechanisms underlying neurodegenerative disorders.