Nature Methods
- 5, 553 - 559 (2008)
Published online: 11 May 2008; | doi:10.1038/nmeth.1212
Real-time imaging of the intracellular glutathione redox potentialMarcus Gutscher1, Anne-Laure Pauleau1, Laurent Marty2, Thorsten Brach2, Guido H Wabnitz3, Yvonne Samstag3, Andreas J Meyer2 & Tobias P Dick11
Redox Regulation Research Group, German Cancer Research Center (DKFZ/A160), Im Neuenheimer Feld 280, D-69120 Heidelberg, Germany. 2
Heidelberg Institute of Plant Sciences, University of Heidelberg, Im Neuenheimer Feld 360, D-69120 Heidelberg, Germany. 3
Institute of Immunology, University of Heidelberg, Im Neuenheimer Feld 305, D-69120 Heidelberg, Germany.
Correspondence should be addressed to Tobias P Dick t.dick@dkfz.de Dynamic analysis of redox-based processes in living cells is now restricted by the lack of appropriate redox biosensors. Conventional redox-sensitive GFPs (roGFPs) are limited by undefined specificity and slow response to changes in redox potential. In this study we demonstrate that the fusion of human glutaredoxin-1 (Grx1) to roGFP2 facilitates specific real-time equilibration between the sensor protein and the glutathione redox couple. The Grx1-roGFP2 fusion protein allowed dynamic live imaging of the glutathione redox potential (E
GSH) in different cellular compartments with high sensitivity and temporal resolution. The biosensor detected nanomolar changes in oxidized glutathione (GSSG) against a backdrop of millimolar reduced glutathione (GSH) on a scale of seconds to minutes. It facilitated the observation of redox changes associated with growth factor availability, cell density, mitochondrial depolarization, respiratory burst activity and immune receptor stimulation.
MORE ARTICLES LIKE THIS These links to content published by NPG are automatically generated.
|
