Imaging the production of singlet oxygen in vivo using a new fluorescent sensor, Singlet Oxygen Sensor Green(R)

Abstract

Singlet oxygen is known to be produced by cells in response to photo-oxidative stresses and wounding. Due to singlet oxygen being highly reactive, it is thought to have a very short half-life in biological systems and, consequently, it is difficult to detect. A new commercially available reagent (singlet oxygen sensor green, SOSG), which is highly selective for singlet oxygen, was applied to a range of biological systems that are known to generate singlet oxygen. Induction of singlet oxygen production by the addition of myoglobin to liposome preparations demonstrated that the singlet oxygen-induced increases in SOSG fluorescence closely followed the increase in the concentration of conjugated dienes, which is stoichiometrically related to singlet oxygen production. Applications of photo-oxidative stresses to diatom species and leaves, which are known to result in the production of singlet oxygen, produced large increases in SOSG fluorescence, as did the addition of 3-(3',4'-dichlorophenyl)1,1-dimethylurea (DCMU) to these systems, which inhibits electron transport in photosystem II and stimulates singlet oxygen production. The conditional fluorescent (flu) mutant of Arabidopsis produces singlet oxygen when exposed to light after a dark period, and this coincided with a large increase in SOSG fluorescence. Wounding of leaves was followed by an increase in SOSG fluorescence, even in the dark. It is concluded that SOSG is a useful in vivo probe for the detection of singlet oxygen.