Super-resolution fluorescence imaging with blink microscopy

Methods Mol Biol. 2013:950:111-29. doi: 10.1007/978-1-62703-137-0_8.

Abstract

Recently, a new approach for super-resolution microscopy has emerged which is based on the successive localization of single molecules. The majority of molecules are prepared to reside in a nonfluorescent dark state, leaving only a few single molecules fluorescing. The single molecules can subsequently be localized on the camera image. Successive localization of all molecules allows reconstruction of a super-resolved image of the labeled structure. A variety of ways for limiting the number of locatable molecules have been developed recently which expand this current field of imaging. Here we describe a super-resolution microscopy method that employs the use of reversible, generic dark states, for example radical ion states. This method requires only a single laser source and can be carried out with many fluorescent dyes, in some cases, even in living cells. We provide a step-by-step procedure for this method, which we have called Blink Microscopy.

MeSH terms

  • Actins / metabolism
  • Cell Adhesion Molecules / metabolism
  • Dendritic Cells / cytology
  • Dendritic Cells / metabolism
  • Fluorescent Dyes / metabolism
  • Humans
  • Image Processing, Computer-Assisted
  • Imaging, Three-Dimensional / methods*
  • Lectins, C-Type / metabolism
  • Microscopy, Fluorescence / instrumentation
  • Microscopy, Fluorescence / methods*
  • Optical Phenomena*
  • Receptors, Cell Surface / metabolism
  • Statistics as Topic

Substances

  • Actins
  • Cell Adhesion Molecules
  • DC-specific ICAM-3 grabbing nonintegrin
  • Fluorescent Dyes
  • Lectins, C-Type
  • Receptors, Cell Surface