524 MYELIN STAINS 



of a chromic salt until they become brown before mordanting them in 

 the copper solution. 



It is not necessary that the mordanting be done in bulk. Max 

 Flesch (Ztschr. wiss. Mikr., iii, 1886, p. 50) prefers (following 

 Lichtheim) to make the sections first, and to mordant them 

 separately. 



Vassale (Riv, sperim. Freniatr., xv, 1889, p. 102) first stains 

 the section in 1 per cent, haematoxylin for three to five minutes, 

 then puts them for three to five minutes into saturated solution 

 of copper acetate, and differentiates as Weigert. 



1057. Weigert's 1891 Method {Deutsche med. Wochenschr., 

 xvii, 1891, p. 1184). The material is hardened in bichromate 

 and imbedded in celloidin (see last §). It is then (according to 

 the latest form of the process {Enzycl. tnik. Technik., 1903, p. 942) ), 

 put for twenty-four hours in a stove into a solution of 2^ parts of 

 chromium fluoride, 5 of copper acetate, and 5 of acetic acid in 

 100 of water.* 



Sections are then made and stained for from four to twenty- 

 four hours at room temperature in a freshly-prepared mixture of 

 9 volumes of (A), a mixture of 7 c.c. of saturated aqueous solution 

 of lithium carbonate with 93 c.c. of water, and 1 volume of (B), 

 a solution of 1 grm. of haematoxylin in 10 c.c. of alcohol (A and B 

 may be kept in stock, but A must not be too old). The sections 

 should be loose ones, and not thicker than 25 /n. They are then 

 washed in several changes of water, and treated with 90 per cent, 

 alcohol, followed by carbol-xylol, or by a mixture of 2 parts of 

 anilin oil with 1 of xylol, then pure xylol and xylol balsam (not 

 chloroform balsam). 



It was, however, found that preparations thus made, without 

 differentiation, did not keep well, and Weigert {Ergebn. Anat., 

 iii, 1894, p. 21) reverted to the practice of differentiating with 

 the borax-ferricyanide mixture. 



Later still {Enzycl. niik. Technik., 1903, p. 942) he employed a 

 stain composed of equal parts of (A), a mixture of 4 c.c. of the 

 officinal Liquor ferri sesquichlorati P. G. with 96 of water, and 

 (B), a mixture of 10 c.c. of a 10 per cent, alcoholic solution of 

 haematoxylin with 90 of 96 per cent, alcohol. The two (A and B) 

 must be mixed immediately before use, and the sections should 

 remain in the stain overnight or longer, then rinsed and 

 differentiated as usual. This has the advantage of demonstrating 

 very fine fibres, and of giving a colourless background. 



For difficult objects the differentiating liquid may be diluted 

 with water, and gives better results than dilute acetic or hydro- 

 chloric acids or the like, which were formerly recommended. 



* Instead of the chromiimi fluoride one may use chrome alum, as 

 Weigert did at one time, and as some still do. But then one must boil, 

 as directed for Weigert's Neuroglia stain. 



