title: Methylene blue- and thiol-based oxygen depletion for super-resolution imaging.
abstract: Anaerobic conditions are often required in solution-based bionanotechnological applications. Efficient oxygen depletion is essential for increasing photostability, optimizing fluorescence signals, and adjusting kinetics of fluorescence intermittency in single-molecule fluorescence spectroscopy/microscopy, particularly for super-resolution imaging techniques. We characterized methylene blue (MB)- and thiol-based redox reactions with the aim of designing an oxygen scavenger system as an alternative to the established enzyme-based oxygen scavenging systems or purging procedures. Redox reactions of the chromophore methylene blue in aqueous solution, commonly visualized in the blue bottle experiment, deplete molecular oxygen as long as a sacrificial reduction component is present in excess concentrations. We demonstrate that methylene blue in combination with reducing compounds such as β-mercaptoethylamine (MEA) can serve as fast and efficient oxygen scavenger. Efficient oxygen scavenging in aqueous solution is also possible with mere β-mercaptoethylamine at mM concentrations. We present kinetic parameters of the relevant reactions, pH-stability of the MB/MEA-oxygen scavenging system, and its application in single-molecule based super-resolution imaging.
authors: Department of Biotechnology & Biophysics, Biocenter, Julius-Maximilians-University Würzburg, Würzburg, Germany.
authors: Philip
authors: P
authors: Schäfer
authors: Sebastian
authors: S
authors: van de Linde
authors: Julian
authors: J
authors: Lehmann
authors: Markus
authors: M
authors: Sauer
authors: Sören
authors: S
authors: Doose
mesh_terms: D006367:HeLa Cells; D006801:Humans; D008751:Methylene Blue / Q000737:chemistry*; D008856:Microscopy, Fluorescence / Q000379:methods; D008870:Microtubules / Q000648:ultrastructure; D010100:Oxygen / Q000737:chemistry*; D013050:Spectrometry, Fluorescence / Q000379:methods; D013438:Sulfhydryl Compounds / Q000737:chemistry*
chemical_list: D013438:Sulfhydryl Compounds; D010100:Oxygen; D008751:Methylene Blue