Dual-color reversible fluorescent carbon dots designed for dynamic monitoring of cellular superoxide anion radicals

Abstract

The superoxide anion radical (O₂˙⁻) represents the primary reactive oxygen species generated in biological systems. Real-time monitoring of its dynamic fluctuations provides valuable insights into disease progression and enables early diagnosis of hepatic ischemia-reperfusion injury (HIRI). In this work, we developed a novel dual-color fluorescent carbon dot (CD) probe through a one-step hydrothermal synthesis for reversible O₂˙⁻ detection. The CDs demonstrated excellent sensitivity, dynamically detecting O₂˙⁻ concentrations ranging from 0 to 60 μM with a detection limit of 0.56 μM. The probe exhibited remarkable reversibility, maintaining stable performance through at least three complete oxidation–reduction cycles following glutathione (GSH) treatment. In practical applications, the CDs achieved 95.2–104% recovery rates when detecting O₂˙⁻ in serum samples. Cellular imaging experiments confirmed the probe's effectiveness in normal hepatocytes (LO2), showing clear reversible responses to O₂˙⁻ fluctuations. Application in a HIRI cell model revealed significant elevation of O₂˙⁻ levels and provided new evidence for its role in HIRI-related signaling pathways. This study not only presents an effective dual-color fluorescent probe for dynamic O₂˙⁻ monitoring but also establishes a versatile synthetic strategy that could be adapted for imaging other biologically relevant molecules in living cells.