3D reconstruction of high-resolution STED microscope images.

Microscopy research and technique (2008-06-03)
Annedore Punge, Silvio O Rizzoli, Reinhard Jahn, Jan Dominik Wildanger, Lars Meyer, Andreas Schönle, Lars Kastrup, Stefan W Hell
ABSTRACT

Tackling biological problems often involves the imaging and localization of cellular structures on the nanometer scale. Although optical super-resolution below 100 nm can be readily attained with stimulated emission depletion (STED) and photoswitching microscopy methods, attaining an axial resolution <100 nm with focused light generally required the use of two lenses in a 4Pi configuration or exceptionally bright photochromic fluorophores. Here, we describe a simple technical solution for 3D nanoscopy of fixed samples: biological specimens are fluorescently labeled, embedded in a polymer resin, cut into thin sections, and then imaged via STED microscopy with nanoscale resolution. This approach allows a 3D image reconstruction with a resolution <80 nm in all directions using available state-of-the art STED microscopes.

MATERIALS
Product Number
Brand
Product Description

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Atto 532 maleimide, BioReagent, suitable for fluorescence, ≥90% (coupling to thiols)
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Atto 532 DOPE, suitable for fluorescence
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Atto 425 maleimide, BioReagent, suitable for fluorescence, ≥70% (coupling to thiols)
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