2. Place in freshly prepared Marchi’s fluid (Müller’s fluid 2 parts, 1 per cent osmic acid solution 1 part) for about 8 days in the incubator at 37°C. The brain requires a longer time. When the mixture loses the osmic acid smell renew it.
3. Wash in running water, 24 hours.
4. Harden in graded alcohols.
5. Imbed in celloidin; cut; dehydrate; clear; mount.
Degenerated nervous tissue (fat) is black: all else brownish gray. Contrast stain in Van Gieson’s, lithium carmine, etc. This method is good for the demonstration of early degenerations.
B. Donaggio’s Methods for Early Degeneration.
Method I—
1. Fix in Müller’s fluid or in 4 per cent potassium-bichromate solution. The tissue may remain in the fluid for any length of time.
2. Transfer directly, without washing, to alcohol. Dehydrate; imbed in celloidin; cut sections 20-30 microns. Place sections in distilled water for a few seconds.
3. Transfer to the following mixture for 10-20 minutes: To 20 per cent solution of ammoniated chloride of tin add an equal amount of 1 per cent aqueous hæmatoxylin. Allow to stand for five days. Keep in the dark, and in a cool place.