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ZFIN ID: ZDB-PUB-171121-12
Correlative Super-resolution and Electron Microscopy to Resolve Protein Localization in Zebrafish Retina
Mateos, J.M., Barmettler, G., Doehner, J., Ojeda Naharros, I., Guhl, B., Neuhauss, S.C.F., Kaech, A., Bachmann-Gagescu, R., Ziegler, U.
Date: 2017
Source: Journal of visualized experiments : JoVE   (129): (Journal)
Registered Authors: Bachmann-Gagescu, Ruxandra, Neuhauss, Stephan
Keywords: none
MeSH Terms:
  • Animals
  • Microscopy, Electron, Scanning/methods*
  • Microscopy, Fluorescence/methods*
  • Retina/diagnostic imaging*
  • Retina/pathology
  • Zebrafish
PubMed: 29155784 Full text @ J. Vis. Exp.
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ABSTRACT
We present a method to investigate the subcellular protein localization in the larval zebrafish retina by combining super-resolution light microscopy and scanning electron microscopy. The sub-diffraction limit resolution capabilities of super-resolution light microscopes allow improving the accuracy of the correlated data. Briefly, 110 nanometer thick cryo-sections are transferred to a silicon wafer and, after immunofluorescence staining, are imaged by super-resolution light microscopy. Subsequently, the sections are preserved in methylcellulose and platinum shadowed prior to imaging in a scanning electron microscope (SEM). The images from these two microscopy modalities are easily merged using tissue landmarks with open source software. Here we describe the adapted method for the larval zebrafish retina. However, this method is also applicable to other types of tissues and organisms. We demonstrate that the complementary information obtained by this correlation is able to resolve the expression of mitochondrial proteins in relation with the membranes and cristae of mitochondria as well as to other compartments of the cell.
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