Live-Cell Imaging of Cytosolic NADH–NAD+ Redox State Using a Genetically Encoded Fluorescent Biosensor

NADH is an essential redox cofactor in numerous metabolic reactions, and the cytosolic NADH–NAD+ redox state is a key parameter in glycolysis. Conventional NADH measurements rely on chemical determination or autofluorescence imaging, which cannot assess NADH specifically in the cytosol of individual live cells. By combining a bacterial NADH-binding protein and a fluorescent protein variant, we have created a genetically encoded fluorescent biosensor of the cytosolic NADH–NAD+ redox state, named Peredox (Hung et al., Cell Metab 14:545–554, 2011). Here, we elaborate on imaging methods and technical considerations of using Peredox to measure cytosolic NADH:NAD+ ratios in individual live cells.
Source: Springer protocols feed by Protein Science - Category: Biochemistry Source Type: news