Single molecule localization microscopy for superresolution

Abstract

In recent years there has been a rash of developments in light microscopies circumventing traditional resolution limits associated with the diffraction of light occurring between the sample and the detector. Collectively, these techniques are referred to as ‘superresolution’ microscopies. One major family of superresolution techniques, variably referred to as PALM, FPALM and STORM, uses temporal control of the excited state of fluorophores to sequentially identify single non-overlapping emitters in time and space. Conventional images of single point emitters are fitted to sub-diffraction-limited areas, and a composite image is reconstructed from position data collected over many thousands of individual imaging frames. This paper provides a brief overview of superresolution microscopy, followed by a detailed discussion of STORM, including practical guidelines for sample preparation designed to help to make the technique more accessible to the non-specialist.