Single-molecule-based super-resolution images in the presence of multiple fluorophores.

ABSTRACT: Several super-resolution techniques exist, yet most require multiple lasers, use either large or weakly emitting fluorophores, or involve chemical manipulation. Here we show a simple technique that exceeds the standard diffraction limit by 5-15× on fixed samples, yet allows the user to localize individual fluorophores from among groups of crowded fluorophores. It relies only on bright, organic fluorophores and a sensitive camera, both of which are commercially available. Super-resolution is achieved by subtracting sequential images to find the fluorophores that photobleach (temporarily or permanently), photoactivate, or bind to the structure of interest in transitioning from one frame to the next. These fluorophores can then be localized via Gaussian fitting with selective frame averaging

SUBMITTER: Simonson PD 

PROVIDER: S-EPMC3212641 | biostudies-literature | 2011 Nov

REPOSITORIES: biostudies-literature