Single-Molecule Localization and Structured Illumination Microscopy of Platelet Proteins

Abstract

Superresolution microscopy has become increasingly widespread over the past 5 years and allows users to image biological processes below the diffraction limit of traditional fluorescence microscopy where resolution is restricted to approximately 250 nm. Superresolution refers to a wide range of techniques which employ different approaches to circumvent the diffraction limit. Two of these approaches, structured illumination microscopy (SIM) and single-molecule localization microscopy (SMLM), which provide a doubling and tenfold increase in resolution respectively, are dominating the field. This is partly because of the insights into biology they offer and partly because of their commercialization by the main microscope manufacturers. This chapter provides background to the two techniques, practical considerations for their use, and protocols for their application to platelet biology.

Keywords: Direct stochastic optical reconstruction microscopy; Platelets; SIM; SMLM; Single-molecule localization microscopy; Structured illumination microscopy; Superresolution; dSTORM.