Morphological Assessment of Intracerebral Hemorrhage - Animal Models of Acute Neurological Injuries: Injury and Mechanistic Assessments - page 589

2. Materials
and Instruments

1. 4% Paraformaldehyde in 0.1 M, pH 7.4 phosphate-buffered
saline (PBS) at 4°C for perfusion.
2. 4% Paraformaldehyde in 0.1 M, pH 7.4 PBS at 4°C.
3. 30% Sucrose in 0.1 M, pH 7.4 PBS at 4°C.
4. O.C.T. compound (Sakura Finetek USA Inc., Torrance, CA).
5. 10% Formalin, neutral buffered (Sigma) for perfusion.
6. 10% Formalin and 70% ethanol for fi xation.
7. Cryostat (Leica ® ) for rats.
8. Thin and sharp blades (Leica ® Disposable Microtome Blades,
Cat. 63065) for pigs before paraffi n embedment.
9. Polycut (Leica ® ) after paraffi ned.
10. Air dryer.
11. Warm incubator.
12. Slide warmer.
13. Microscope equipped with digital camera for rats and pigs.
14. Digital camera, ruler, and Blue plastic that provides a good
contrast to red and white for pigs.
15. NIH ImageJ software (NIH, Bethsda, MD).

3. Procedures

3.1. Brain Sample
Preparation

For rats,
Perfuse rat brains intracardiacally with cold 4% paraformalde-

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hyde in 0.1 M, pH 7.4 PBS.
Fix brains with 4% paraformaldehyde at 4°C overnight.

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Dehydrate brains with 30% sucrose in 0.1 M, pH 7.4 PBS for

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3-4 days at 4°C.
Embed brains in 2/3 of 30% sucrose and 1/3 of O.C.T. com-

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pound on dry ice and store the brains in −80°C freezer.
For pigs,
Perfuse pig brains intracardiacally with 10% formalin.

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Fix the brain in 10% formalin solution at room temperature for

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1 month and move the brain to 75% ethanol solution before
brain sectioning.