8o Neurological Technique 



For convenience in sectioning, the piece of nerve or spinal 

 cord should be held straightened out while fixing and harden- 

 ing. This can be done by tying it at the ends to a strip of 

 glass small enough to go into the vessel of fixing fluid. 



2. Cut out the region from which sections are desired and 

 wash in several changes of distilled water, 4-8 hours, 



3. Dehydrate with alcohol, imbed in celloidin, and section as 

 directed in X, 3, 4, and 5, p. 64. Make sections about 15 /* 

 thick. If serial sections are desired, proceed as in XI, p. 72. 



4. To stain. — Withdraw 70 per cent, alcohol from the sec- 

 tions and replace it with 50 per cent, for about 5 minutes. Then 

 replace 50 per cent, alcohol with the aniline-blue solution (B) 

 for 20-60 minutes. 



5. To differentiate. — {a) Rinse in 2 or 3 changes of distilled 

 water for not more than 5 minutes. The sections are now a 

 blue-black. 



((5) Then transfer the sections, one at a time, to a shallow 

 dish of 95 per cent, alcohol, to every 30 cc. of which there has 

 been added 2 cc. of the alcoholic i per cent, caustic potash solu- 

 tion (D). Agitate the fluid about the section till it acquires a 

 light brownish-red color. This usually takes from 1-3 minutes. 



If it takes more than 5 minutes, either the section is too 

 thick or a few more drops of the alkaline alcohol are necessary. 



6. Wash in several changes of distilled water for 10-15 min- 

 utes. 



Here the sections should become a clear bright blue. If 

 any appear black-blue, return them to the differentiating fluid 

 for a few minutes longer, and then wash as before. 



7. To counterstain. — Place the thoroughly washed sections in 

 a small amount of the saturated aqueous safranin solution (c) , 

 to which an equal amount of distilled water has been added, for 

 15-30 minutes. 



8. To clear and mount, quickly wash out the excess of* 

 safranin in 97 per cent, alcohol (8-10 seconds in about 50 cc. 

 in a shallow dish), and then, to clear, place the sections in oil 

 of origanum or carbol-xylol (XII, C, p. 75), 10-15 minutes. 



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