Microtome

🔬 Microtome Protocol

This protocol outlines the materials and steps for preparing and slicing tissue using a microtome/cryostat.

📋 Materials

• Microtome
• Dry Ice
• Paper towel
• Brain Slice
• Razor Blade
• Paint brush
• 12 Well plate
• Larger well plate
• PBS (Phosphate-Buffered Saline)
• Mounting Medium (OCT)
• Aluminum Foil
• Styrofoam box

📝 Protocol

1. Pour the sucrose into the sink and put the brain on aluminum foil in the dry ice in a Styrofoam box. Don't have the brain touch the dry ice. Dry the brain with a paper towel.
2. Put dry ice around the microtome tools to freeze the surface.
3. Using the OCT, add a generous amount of the mounting medium in the middle of the microtome.
4. Place the brain on the mounting medium with the most posterior segment of the brain lying on the mounting medium. Try to ensure it is level. Use more OCT to secure the brain. (If walking away, you can use Styrofoam to insulate it. This is to keep the anterior brain cold.
5. As the brain is cooling, put 1x PBS in a 12 deep well plate and a larger well plate.
6. Before starting to cut, make sure the blade is clean with ethanol and wet with PBS.
7. Now move the blade forward and back to get slices, making sure the blade goes completely back to its neutral spot to allow the microtome to adjust.
8. To remove brain slices, use the paint brush with PBS and put into the 12 well plate, separating based on brain location.
9. Once done, remove the brain from the mounting media with the razor blade.
10. Clean area with ethanol.