METHOD V. 



KUPFFER'S METHOD. 



A method for the study of the medullated axone — the myelin 

 sheath, the axis cylinder, nodes of. Ranvier, etc. 



SPECIAL REAGENTS REQUIRED. 



A. Fixing agent (0.5 per cent, osmic acid). — 



Osmic acid crystals (in sealed tube), 0.5 g. 



Pure distilled water, 100 cc. 

 The bottle must be chemically clean. For directions as to making osmic 

 acid solutions see IV, A, note, p. 31. A stronger solution may be reduced 

 by adding the necessary amount of distilled water. 



B. Stain (saturated aqueous acid fuchsin). — 



Fuchsin S {Sdurefucksin, Griibler), 1.5 g. 

 Distilled water, 100 cc. 



PROCEDURE. 



I. To fix. — Carefully lay bare a small nerve and, without 

 stretching, gently force under it the end of a strip of cardboard 

 about ^2 inch wide. Cut so that the resulting piece of nerve 

 will be 2 or 3 millimeters longer than the cardboard is wide. 

 The nerve will adhere to the cardboard, and thus, when cut at 

 each side, a piece is obtained held straight, but, at the same 

 time, under very nearly its normal tension. The end of the 

 cardboard holding the bit of nerve is then clipped off into a 

 clean vial containing 10-15 cc of the 0.5 per cent, osmic acid 

 solution (A), and allowed to remain in the fluid for 4-24 

 hours. 



The nerve should be quite small to insure the best penetration 

 of the osmic acid. The spinal nerve-roots afford an opportunity 

 to get nerve-bundles of good penetrable size, and also those from 

 the lumbar region can be easily removed, since they lie free in 

 the vertebral canal. 



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