76 Neurological Technique 



only colored a light yellow, due to the bichromate in the Miil- 

 ler's fluid. The other ingredients of the Muller's fluid only serve 

 to improve its fixing properties. 



A critical discussion of Marchi's method and its modifications 

 up to that date may be found in Ergebnisse der Anatomie (Merkel 

 and Bonnet), Bd. VII, 1897. 



PROCEDURE. 



1. The lesion or operation leading to the degeneration of 

 the fibers the course of which is to be studied should take place 

 2-4 weeks before the animal is killed and the tissue removed 

 for fixation. This time may vary for different species of ani- 

 mals. The optimum time is when the degenerative changes 

 upon which the method depends are at their height. In human 

 pathology it has been found that results may be obtained even 

 after the processes of degeneration have been going on for sev- 

 eral months. 



It is not necessary that the tissues be absolutely fresh. It 

 has even been recommended that the animal be killed and 

 allowed to lie in a cool place 24 hours before the tissues are 

 removed. This, however, is now considered by no means essen- 

 tial to the method. 



2. Remove pieces of tissue (not thicker than 10 mm.) from 

 the required locality and place in 20-30 volumes of Miiller's 

 fluid (A), and set aside, preferably in the dark, for 10-15 days. 



Change the fluid after the first and third days. The time 

 may be reduced to 8 days by keeping the temperature of the 

 fluid at 35" or 40° C. 



3. If the material is from the central nervous system, cut 

 the pieces into sections not thicker than 4 mm. (the thinner the 

 better), and place them in 10-15 volumes of the osmic acid mix- 

 ture (B), and keep in the dark for 4-6 days. For peripheral 

 nerves 2-4 days in the fluid is sufficient. 



Change the fluid after the first and third day. The mixture 

 should be freshly made each time by adding the osmic acid to 

 the Muller's fluid. 



4. Wash in several changes of distilled water for 8-24 hours. 



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