Singlet oxygen imaging using fluorescent probe Singlet Oxygen Sensor Green in photosynthetic organisms

Abstract

Formation of singlet oxygen (1O2) was reported to accompany light stress in plants, contributing to cell signaling or oxidative damage. So far, Singlet Oxygen Sensor Green (SOSG) has been the only commercialized fluorescent probe for 1O2 imaging though it suffers from several limitations (unequal penetration and photosensitization) that need to be carefully considered to avoid misinterpretation of the analysed data. Herein, we present results of a comprehensive study focused on the appropriateness of SOSG for 1O2 imaging in three model photosynthetic organisms, unicellular cyanobacteria Synechocystis sp. PCC 6803, unicellular green alga Chlamydomonas reinhardtii and higher plant Arabidopsis thaliana. Penetration of SOSG differs in both unicellular organisms; while it is rather convenient for Chlamydomonas it is restricted by the presence of mucoid sheath of Synechocystis, which penetrability might be improved by mild heating. In Arabidopsis, SOSG penetration is limited due to tissue complexity which can be increased by pressure infiltration using a shut syringe. Photosensitization of SOSG and SOSG endoperoxide formed by its interaction with 1O2 might be prevented by illumination of samples by a red light. When measured under controlled conditions given above, SOSG might serve as specific probe for detection of intracellular 1O2 formation in photosynthetic organisms.

Anticancer Photodynamic Therapy Properties of Sulfur‐Doped Graphene Quantum Dot and Methylene Blue Preparations in MCF‐7 Breast Cancer Cell Culture

“…Singlet oxygen in MCF‐7 cells was detected using epifluorescent microscopy to measure the fluorescence of SOSG dye following a procedure in the literature with slight modification. Two 24‐well plates were seeded with 50 000 MCF‐7 cells in four wells per plate in media with either 23 μ g mL −1 MB, 20 μ g mL −1 GQD, volumetric 1:1 GQD:MB or media only in each well.…”

Oxygen and ROS in Photosynthesis

“…The formation of the excited singlet state 1 P680* is followed by the formation of the radical pair [190], histidine [191][192][193] and Singlet Oxygen Sensor Green [190,194,195]. The 1270 nm luminescence has been used to measure 1 O 2 generation by isolated RC complexes [171,196].…”

Adaptation to Photooxidative Stress: Common and Special Strategies of the Alphaproteobacteria Rhodobacter sphaeroides and Rhodobacter capsulatus

“…To see whether the different Crt:BChl a ratios observed in the two Rhodobacter species under photooxidative stress conditions ( Figure 2D) would affect 1 O 2 levels, the fluorescent probe Singlet Oxygen Sensor Green (SOSG) was used for in vivo 1 O 2 measurements. Since white light itself can affect SOSG fluorescence, red light was used for illumination [62]. In cell-free reactions, the combination of oxygen, red light, and methylene blue caused enhanced SOSG fluorescence due to photosensitized formation of 1 O 2 ( Figure S4).…”