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Title:
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Aldehyde-stabilized cryopreservation
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Author: |
R. McIntyre, G.M. Fahy
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Citation: |
Cryobiology 71 (2015) 448-458
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Abstract: |
We describe here a new cryobiological and neurobiological technique, aldehyde-stabilized cryopreservation
(ASC), which demonstrates the relevance and utility of advanced cryopreservation science for the
neurobiological research community. ASC is a new brain-banking technique designed to facilitate
neuroanatomic research such as connectomics research, and has the unique ability to combine stable
long term ice-free sample storage with excellent anatomical resolution. To demonstrate the feasibility of
ASC, we perfuse-fixed rabbit and pig brains with a glutaraldehyde-based fixative, then slowly perfused
increasing concentrations of ethylene glycol over several hours in a manner similar to techniques used
for whole organ cryopreservation. Once 65% w/v ethylene glycol was reached, we vitrified brains
at -135°C for indefinite long-term storage. Vitrified brains were rewarmed and the cryoprotectant
removed either by perfusion or gradual diffusion from brain slices. We evaluated ASC-processed brains
by electron microscopy of multiple regions across the whole brain and by Focused Ion Beam Milling and
Scanning Electron Microscopy (FIB-SEM) imaging of selected brain volumes. Preservation was uniformly
excellent: processes were easily traceable and synapses were crisp in both species. Aldehyde-stabilized
cryopreservation has many advantages over other brain-banking techniques: chemicals are delivered via
perfusion, which enables easy scaling to brains of any size; vitrification ensures that the ultrastructure of
the brain will not degrade even over very long storage times; and the cryoprotectant can be removed,
yielding a perfusable aldehyde-preserved brain which is suitable for a wide variety of brain assays.
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Article Link |
Http://www.21CMPublications.com/PubFiles/102/2015McIntyreFahyASC.pdf
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