Rapid decapitation and brain cryopreservation

1. Guillotine used for rapid decapitation on anesthetized (isoflurane) rats
2. Quickly but carefully remove the brain from the skull
3. Brain placed on moistened (PBS) filter paper on upside-down beaker in a cooler of wet ice 
4. Brain rinsed with PBS to remove blood
5. Use an extra piece of filter paper to remove excess liquid
6. Keep the brain whole (coronal) or hemisect brain (sagittal) and with a spatula closer to the surface of the liquid, drop into SUPER COLD (on dry ice) 2-methyl butane
7. The brain should be fully covered by 2-methyl butane; leave in liquid until the surface looks porcelain (~60 secs)
8. With cold tweezers, remove the brain from 2MB and place it on a piece of dry ice until the surface appears dry
9. Put in a pre-labeled tube that can be cooled on dry ice- store long term at -80C

Dry Ice can be found in MBB 1.122, purchase using the following:

https://docs.google.com/forms/d/e/1FAIpQLSfxucgcCOdEHFO-zT832qu_Fv3GEzuxYE48d_-A3a97L6bosg/viewform
acct number: 26-1679-7650