Dec 14, 2023

Public workspaceCryoprotection of Mouse Brain

DOI
dx.doi.org/10.17504/protocols.io.yxmvm358ol3p/v1
  • 1University of Alabama at Birmingham, Department of Neurology, Center for Neurodegeneration and Experimental Therapeutics
Jhodi Webster
Open access
DOI: dx.doi.org/10.17504/protocols.io.yxmvm358ol3p/v1
Protocol Citation: Asta Zane, Nicole J Corbin-Stein, Gabrielle Childers, Jhodi Webster, Vickie Yang, Woong-Jai Won, Rajesh Gupta, Ashley Harms 2023. Cryoprotection of Mouse Brain. protocols.io https://dx.doi.org/10.17504/protocols.io.yxmvm358ol3p/v1
Manuscript citation:
Williams GP, Schonhoff AM, Jurkuvenaite A, Gallups NJ, Standaert DG, Harms AS. CD4 T cells mediate brain inflammation and neurodegeneration in a mouse model of Parkinson's disease. Brain. 2021 Aug 17;144(7):2047-2059. doi: 10.1093/brain/awab103. PMID: 33704423; PMCID: PMC8370411.

Schonhoff, A.M., Figge, D.A., Williams, G.P. et al. Border-associated macrophages mediate the neuroinflammatory response in an alpha-synuclein model of Parkinson disease. Nat Commun 14, 3754 (2023). https://doi.org/10.1038/s41467-023-39060-w
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: November 03, 2023
Last Modified: May 31, 2024
Protocol Integer ID: 90384
Keywords: ASAPCRN
Funders Acknowledgement:
Co-pathologies Drive Neuroinflammation and Progression in PD
Grant ID: ASAP-021030
Abstract
This protocol allows for accurate cryoprotection of mouse brain, post-perfusions, to be used for histology. The methods utilize another PFA fixation step followed by sucrose incubation.
Materials
  •       Cold Paraformaldehyde (PFA) solution 4% in PBS 
  •       30% sucrose in phosphate buffered saline (PBS). To make it, mix 30g of sucrose with PBS solution to a total volume of 100ml.
  •       Dry ice
  •       Foil
  •       15 ml conical tube
PROCEDURE
PROCEDURE
Transfer the brain into 5-10ml of 4% PFA solution in PBS for 2 hours at room temperature in a 15 ml conical tube.
Transfer the brain into 30% sucrose solution in PBS, wait until it sinks to the bottom for 48-72 hours at 4°C. 
Freeze brain on dry ice and store at minus -80 °C till sectioning.   
To freeze the brain cut foil into pieces that will be labeled accordingly. Lay the foil on the top of the dry ice. Put the brain on the foil. Let the brain to freeze. It should change its color to white, and become hard. Wrap the foils edges. Put samples in the box, label the box with your project, and put it to minus -80 °C.