Nature Methods: Live-cell dSTORM with SNAP-tag fusion proteins.
10.01.2011Teresa Klein, Anna Löschberger, Sven Proppert, Steve Wolter, Sebastian van de Linde & Markus Sauer Nature Methods 8, 7–9 (2011) | doi:10.1038/nmeth0111-7b Published online 29 December 2010
Abstract
In the September 2010 issue of Nature Methods we demonstrated live-cell direct stochastic optical reconstruction microscopy (dSTORM) of histone H2B proteins using a trimethoprim chemical tag (TMP tag) for genetic encoding with photostable standard fluorophores. The method takes advantage of the fact that cells contain the reducing thiol glutathione—a cysteine-containing tripeptide—at millimolar concentrations, which enables reversible photoswitching of synthetic organic fluorophores. The generality of the method can be easily understood considering that most Alexa Fluors (Invitrogen) and Atto fluorophores (ATTO-TEC) belong to the class of rhodamine and oxazine derivatives that have similar redox properties, that is, the triplet state of rhodamine and oxazine fluorophores is reduced by thiols such as glutathione.
