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024 7 _ |2 pmid
|a pmid:22301777
024 7 _ |2 DOI
|a 10.1038/nprot.2011.449
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041 _ _ |a eng
082 _ _ |a 610
084 _ _ |2 WoS
|a Biochemical Research Methods
100 1 _ |a Marx, M.
|b 0
|u FZJ
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245 _ _ |a Improved biocytin labeling and neuronal 3D reconstruction
260 _ _ |a Basingstoke
|b Nature Publishing Group
|c 2012
300 _ _ |a 394 - 407
336 7 _ |a Journal Article
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440 _ 0 |a Nature Protocols
|0 24802
|v 7
500 _ _ |a This work was supported by the Deutsche Forschungsgemeinschaft (DFG; Research Group BaCoFun), the Helmholtz Association and the Helmholtz Alliance for Systems Biology. We thank A. Rollenhagen for help with the EM protocol and T. Abel for critically reading the manuscript.
520 _ _ |a In this report, we describe a reliable protocol for biocytin labeling of neuronal tissue and diaminobenzidine (DAB)-based processing of brain slices. We describe how to embed tissues in different media and how to subsequently histochemically label the tissues for light or electron microscopic examination. We provide a detailed dehydration and embedding protocol using Eukitt that avoids the common problem of tissue distortion and therefore prevents fading of cytoarchitectural features (in particular, lamination) of brain tissue; as a result, additional labeling methods (such as cytochrome oxidase staining) become unnecessary. In addition, we provide correction factors for tissue shrinkage in all spatial dimensions so that a realistic neuronal morphology can be obtained from slice preparations. Such corrections were hitherto difficult to calculate because embedding in viscous media resulted in highly nonlinear tissue deformation. Fixation, immunocytochemistry and embedding procedures for light microscopy (LM) can be completed within 42-48 h. Subsequent reconstructions and morphological analyses take an additional 24 h or more.
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650 _ 2 |2 MeSH
|a Animals
650 _ 2 |2 MeSH
|a Brain: cytology
650 _ 2 |2 MeSH
|a Brain: ultrastructure
650 _ 2 |2 MeSH
|a Imaging, Three-Dimensional: methods
650 _ 2 |2 MeSH
|a Lysine: analogs & derivatives
650 _ 2 |2 MeSH
|a Mice
650 _ 2 |2 MeSH
|a Microtomy: methods
650 _ 2 |2 MeSH
|a Neurons: ultrastructure
650 _ 2 |2 MeSH
|a Osmium Tetroxide
650 _ 2 |2 MeSH
|a Rats
650 _ 2 |2 MeSH
|a Staining and Labeling: methods
650 _ 7 |0 20816-12-0
|2 NLM Chemicals
|a Osmium Tetroxide
650 _ 7 |0 56-87-1
|2 NLM Chemicals
|a Lysine
650 _ 7 |0 576-19-2
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|a biocytin
650 _ 7 |a J
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700 1 _ |a Günter, R.H.
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700 1 _ |a Hucko, W.
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700 1 _ |a Radnikow, G.
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700 1 _ |a Feldmeyer, D.
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|0 P:(DE-Juel1)131680
773 _ _ |a 10.1038/nprot.2011.449
|g Vol. 7, p. 394 - 407
|p 394 - 407
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|t Nature protocols
|v 7
|y 2012
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856 7 _ |u http://dx.doi.org/10.1038/nprot.2011.449
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