Single-molecule localization microscopy comes of age: From concepts to biological impact

Abstract

Superresolution microscopy by single-molecule photoactivation or photoswitching and position determination (localization microscopy) has the potential to fundamentally revolutionize our understanding of how cells function at a molecular level. Single-molecule localization microscopy (SMLM) can provide single-molecule information about the distribution and even the absolute numbers of proteins in subcellular compartments. During this webinar, our speakers will introduce the basic requirements of SMLM and its potential use for quantitative molecular imaging. Obstacles and how to bypass them will be discussed, as well as the advantages of direct stochastic optical reconstruction microscopy (dSTORM) for quantitative imaging of proteins. Furthermore, the influence of different environmental and artificial variables on the survival of different cell lines will be demonstrated. The speakers will present data on different image capture technologies for SMLM, focusing on electron multiplying charge-coupled device (EMCCD) and scientific complementary metal-oxide semiconductor (sCMOS) cameras, discussing frame rates and the need for sensitive detectors to harvest the comparatively low light levels associated with single-molecule fluorescence emission. Information on how to optimize your camera setup for superresolution imaging will also be provided.