Indirect imaging of singlet oxygen generation from a single cell

Singlet oxygen (¹O₂) can be generated in a living cell upon focused laser irradiation of the intracellular photosensitizers. ¹O₂ lifetime in the living cells is shortened by the reactions with cellular molecules, and thus the ¹O₂ diffusion in a single cell has attracted much attention. In this study, ¹O₂ gen-eration from the plasma membrane-targeted protoporphyrin IX (PpIX) and nuclear-targeted meso-Tetra (N-methyl-4-pyridyl) porphine tetra tosylate (TMPyP) in human nasopharyngeal carcinoma CNE2 cells was indirectly imaged by using a fluorescence probe Singlet Oxygen Sensor Green agent (SOSG), respectively. The confocal images indicate that the green fluorescence of SOSG in the vicinity of the PpIX-sensitized cells was dramatically enhanced with the increase of the irradiation time and intracellular PpIX, while there is no significant enhancement for the unsensitized and TMPyP-sensitized cells. The obtained results suggest that the ¹O₂ generated from the plasma membrane-targeted PpIX in the CNE2 cells can escape into the extracellular medium and react with the SOSG to produce SOSG endoperoxides (SOSG-EP). Moreover, the fluorescence enhancement of SOSG mainly depends on the subcellular localization and intracellular uptake of the photosensitizers. Depending on the site of ¹O₂ generation, ¹O₂ generated in the plasma membrane can escape from the cell interior into the extracellular environment, while the ¹O₂ generated in the nucleus cannot. Our findings indicate that SOSG holds great promise for the indirect imaging of the ¹O₂ that can escape from single intact living cells.