Three-dimensional, Tomographic Super-Resolution Fluorescence Imaging of Serially Sectioned Thick Samples

PLoS One. 2012;7(5):e38098. doi: 10.1371/journal.pone.0038098. Epub 2012 May 25.

Abstract

Three-dimensional fluorescence imaging of thick tissue samples with near-molecular resolution remains a fundamental challenge in the life sciences. To tackle this, we developed tomoSTORM, an approach combining single-molecule localization-based super-resolution microscopy with array tomography of structurally intact brain tissue. Consecutive sections organized in a ribbon were serially imaged with a lateral resolution of 28 nm and an axial resolution of 40 nm in tissue volumes of up to 50 µm×50 µm×2.5 µm. Using targeted expression of membrane bound (m)GFP and immunohistochemistry at the calyx of Held, a model synapse for central glutamatergic neurotransmission, we delineated the course of the membrane and fine-structure of mitochondria. This method allows multiplexed super-resolution imaging in large tissue volumes with a resolution three orders of magnitude better than confocal microscopy.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anatomy, Cross-Sectional / methods*
  • Animals
  • Brain / cytology
  • Brain / ultrastructure
  • Imaging, Three-Dimensional / methods*
  • Microscopy, Fluorescence / methods*
  • Mitochondria / ultrastructure
  • Rats
  • Rats, Sprague-Dawley
  • Synapses / ultrastructure
  • Tomography / methods*