Point by Point: An Introductory Guide to Sample Preparation for Single-Molecule, Super-Resolution Fluorescence Microscopy.
Single-molecule, localization-based, super resolution microscopy is able to reveal detailed subcellular structures and protein distributions below the classical ∼250-nm diffraction limit of light, but utilizing this technique effectively requires a combination of careful sample preparation, data acquisition, and data analysis, which can be daunting to novice researchers. In this protocol, detailed instructions on preparation of robust reference samples for super-resolution microscopy, including the cytoskeleton (microtubules), membrane-bound organelles (peroxisomes), and scaffold proteins (clathrin), are provided. These samples also constitute a representative subset of imaging targets of interest to biological researchers and highlight the differences and similarities in sample preparation.
Department of Chemistry, University of Washington, Seattle, Washington.
Aaron R
AR
Halpern
Department of Chemistry, University of Washington, Seattle, Washington.
Marco D
MD
Howard
Department of Chemistry, University of Washington, Seattle, Washington.|Department of Physiology and Biophysics, University of Washington, Seattle, Washington.
Joshua C
JC
Vaughan
D053000:Analytic Sample Preparation Methods; D000818:Animals; D002460:Cell Line; D002966:Clathrin / Q000648:ultrastructure; D008856:Microscopy, Fluorescence / Q000379:methods*; D008870:Microtubules / Q000648:ultrastructure; D020675:Peroxisomes / Q000648:ultrastructure
D002966:Clathrin
PALM; STORM; clathrin; microtubules; peroxisomes; super-resolution microscopy