STAINS, ETC. 



1394 



STAINING OF NERVE-TISSUE 



hardened for 15 minutes in 70 per cent, alcohol, cut, 

 and mounted in the usual way. Golgi's Sublimate 

 Method. Harden the tissue for from 15 to 20 days in 

 Midler's fluid, and pass it directly into a 0.25 or 0.5 

 per cent, solution of mercuric chlorid, which should be 

 renewed until it no longer turns yellow. After at least 

 10 days' immersion in this solution, sections are cut on a 

 freezing microtome, washed thoroughly in water, dehy- 

 drated, cleared, and mounted. The nerve-cells, with 

 their processes and nuclei, are brought out prominently. 

 Pal's modification of this method consists in after-treat- 

 ment of the sections with a weak solution of sodium 

 sulphid. Hoyer's Method. For corneal nerves. Im- 

 merse corneas for from *^ to 5 hours, according to size, 

 in an acidulated o 5 per cent, solution of potassio-gold 

 chlorid. To demonstrate the intra-epithelial ramifications 

 of nerves, reduce for from 16 to 24 hours by exposure in 

 distilled water containing 2 drops of pyrogallic-acid de- 

 veloping solution, such as is used in photography ; or, 

 instead, the corneoe may be placed in a warm, concen- 

 trated solution of tartaric acid at the temperature of an 

 incubating oven until the gold is reduced. Isolated 

 Neuroglia Cells. Make an interstitial injection of 

 I per cent, osmic acid into the white matter of the 

 spinal cord. Tease a piece, and stain it with picro- 

 carmin. Jakimovitch's Method. Used to demon- 

 strate Frohmann's lines and Ranvier's crosses. Place 

 small pieces of nerve, in the dark, in a I per cent, silver- 

 nitrate solution ; continue the immersion for 48 hours, 

 and renew the solution frequently. Wash in water, and 

 expose to light for from 5 to 7 days in I part each of 

 formic acid and amyl-alcohol in 100 parts of water. 

 Tease, and mount in glycerin. Joseph's (M.) 

 Method. Used to demonstrate Frohmann's lines and 

 Ranvier's crosses. Place the fresh nerve in 1 per cent, 

 silver nitrate and 10 per cent, nitric acid mixture 

 diluted with water. After several hours, transfer to a 

 weak solution of potassium bichromate, and thence 

 pass through solutions of increasing concentration until 

 it is hardened. It may then be teased and mounted. 

 Kaiser's Method. Stain celloidin sections of spinal 

 cord in a solution of I part naphthylamin-brown 

 (Griibler), alcohol 100 parts, water 200 parts. Wash 

 with alcohol and clear with origanum-oil. Chromo- 

 philous ganglion-cells appear dark-brown ; chromo- 

 phobous cells, light on a dark ground. Korybutt- 

 Daszkiewicz's Method. For the study of the cen- 

 tral nervous system. Secure sections to the slide by 

 means of distilled water. Stain for I minute in Bohm- 

 er's hematoxylin, and wash in a I per cent, solution of 

 alum and distilled water ; stain for I minute in a 1 per 

 cent, aqueous solution of nigrosin, and wash in water ; 

 stain for from 15 to 20 seconds in a 0.5 per cent, 

 alcoholic, watery solution of eosin, and wash for a few 

 minutes in distilled water ; transfer to absolute alcohol ; 

 stain for 20 minutes in a 0.5 per cent, alcoholic, watery 

 solution of safranin ; wash well in alcohol ; clear, but 

 not in clove-oil, and mount in balsam. Kupffer's 

 Method. A nerve is stretched on a cork, and treated 

 for 24 hours with 0.5 per cent, osmic acid; then 

 washed in water for 2 hours, and stained for from 24 

 to 28 hours in a saturated, aqueous solution of acid 

 fuchsin. After this it is washed out for from 6 to 1 2 

 hours (not more in any case) in absolute alcohol, and 

 cleared in clove-oil, embedded in paraffin, and cut. 

 The axis-cylinder appears as a bundle of red fibrils 

 floating in an albuminous liquid. Magini's Method. 

 Used to demonstrate the finer stmcturc of ganglion-cells 

 and their processes. Harden cubes of from 2 to 3 cm. 

 for from 2 to 3 months in Miiller's fluid, wash well 

 with distilled water, and bring for 10 days into from a 

 0.5 to a 1 per cent, solution of zinc chlorid, which 



should be changed daily, until it does not becom 

 yellower than bichromate solution. Section, was 

 quickly with alcohol, clear partially with creasc-U 

 and mount in dammar. Marchi's Method. Usaj t 

 demonstrate early degeneration of nerves, prior t 

 sclerosis. After hardening in Miiller's fluid, plac 

 the tissue in a large quantity of a mixture of Mil 

 ler's fluid 2 parts, I per cent, osmic acid 1 pari 

 The degenerated fibers are stained black, th 

 normal are yellow or uncolored. Martinotti' 

 Method. Stain for 2 or 3 hours or clays in a sal 

 urated solution of nigrosin in a saturated solution 

 picric acid in alcohol ; wash out in a mixture of 1 pai 

 formic acid and 2 parts alcohol, until the gray sul: 

 stance appears to the unaided eye differentiated froi 

 the white. This method is of value in the stud 

 of pathologic tissue. May's Methods. 1. Forpeript 

 eral nerves and ganglia, macerate pieces of muscle i 

 0.5 per cent, arsenic acid, and when swollen place fc 

 20 minutes in solution of 1 per cent, potassio-gol 

 chlorid 4 c.c, 2 per cent, osmic acid I c.c, 0.5 pe 

 cent, arsenic acid 20 c.c. Then wash in water, an 

 expose to sunlight for 3 hours in a I per cent, soli 

 tion of arsenic acid, kept at 45 C. in a water-batl 

 Clear in a mixture of glycerin 40 c.c. , water 20 c.c 

 25 per cent, hydrochloric acid I c.c. 2. Treat asinaj 

 piece of muscle for 12 hours with water containing j 

 per cent, of glacial acetic acid, and transfer to a fresh 

 made solution of 0.5 per cent, potassio-gold chlorid I 

 c.c, 2 per cent, osmic acid, 1 c.c, 2 per cent, glacil 

 acetic acid 50 c.c. After from 2 to 3 hours bring in 

 acidulated glycerin, and when the tissue is transparer 

 examine in glycerin or Farrant's solution. Modi j 

 cations of Weigert's Method. 1. Benda s MoX 

 fication. Small pieces of nervous tissue are placed 

 3 days or more in a saturated solution of picric ac I 

 washed in water, and the hardening continued in aid 

 hoi. Embedding in paraffin is best. Thin sections :| 

 placed in a concentrated solution of iron sulphate, wasH 

 repeatedly, and put into a I per cent, solution of hej ■ 

 atoxylin until deep-black in color (about 10 minutel 

 They are bleached in a solution of chromic acid, 1 : 2C : 

 washed, dehydrated, and mounted. The fibers and } . 

 intimate structure of the cells are well brought c 

 2. Berkley 's Modification. Small pieces are fixecT 

 Flemming's solution for from 24 to 30 hours, at a t< 

 peratureof 25 C, then transferred directly into abso'f -;■ 

 alcohol, which is changed twice during the following I 

 hours, and then placed from 12 to 24 hours in celloiJ 

 Thin sections should be cut, washed in water, and I- 

 into a saturated solution of copper acetate over night 1 

 a covered vessel. If necessary to be rapidly d 

 sections may be heated over a water-bath to between r 

 and 40 C. for 25 minutes, and then allowed to cj. - ' 

 After washing in water the sections are stained I 

 hematoxylin solution, prepared as follows: 50 c.c 1 

 water are boiled in a flask and 2 c.c. of a satum 

 tion of lithium carbonate added ; boil a little longer, | 

 add 1.5 or 2 c.c. of a 10 per cent, solution of hen 

 in absolute alcohol. The flask is then shaken. 

 and allowed to cool. The solution improve- 

 or two, but may be used at once. After staining 1 

 sections are washed and put into Weigert's ■ 

 potassium ferrocyanid solution, which may i 

 one-third. The decolorizing ought to be com 

 from 1 to 3 minutes. Then wash several times in w I 

 then in alcohol, and mount in xylol-balsam. 

 medullated nerves appear blue-black, the glia 

 yellowish, the nerve-cells unstained. 3. Breg 

 i fication. Mordant sections for from 10 to 15 minulB" 

 i< mixture of 15 c.c. of 90 per cent, alcohol and froi'! 10 

 7 c.c. of a saturated aqueous solution of iieutr; 



