STAINS, ETC. 



1396 



STAINING OF NERVE-TISSUE 



substance. Transfer to a o. I per cent gold-chlorid solu- 

 tion until the gray matter shows a violet tinge, wash 

 carefully in water, place in absolute alcohol. When the 

 rosy- violet of the gray substance forms a distinct contrast 

 to the red medullary substance carry to clove-oil, then 

 to xylol or toluol, and mount in balsam. Nissl's 

 Method. Treat bichromate objects with 95 per cent, 

 alcohol ; then stain for 72 hours in an aqueous solution 

 of Congo red, 5 : 400 ; wash out for from 5 to IO minutes 

 in 95 per cent, alcohol ; treat for 6 hours with a 3 per 

 cent, solution of nitric acid in alcohol ; dehydrate for 5 

 minutes in alcohol, clear in clove-oil and mount in 

 balsam. A specific stain for axis-cylinders. Paladino's 

 Method. For axis-cylinders. Pieces not more than 

 from 5 to 8 mm. in thickness, hardened in potassium bi- 

 chromate , chromic acid, or corrosive sublimate, are put for 

 two days into a large quantity (200 c. c. for each piece) of 

 a O.I per cent, aqueous solution of palladium chlorid. 

 Then place them for 24 hours in a I per cent, solution of 

 potassium iodid, using a relatively small volume, or the 

 palladium iodid formed in the tissues may be extracted. 

 After from 1 to 2 hours, dehydrate and embed in paraffin 

 by the chloroform method. Medulla, axis-cylinders, and 

 cell-processes are stained brown. Platner's Method. 

 Small, fresh nerves are fixed and hardened for several 

 days in I part of ferric-chlorid solution and 3 or 4 of 

 water or alcohol, washed out in water or alcohol until the 

 washings no longer give the reaction for iron with potas- 

 sium rhodanid, and stained for several days or weeks in a 

 concentrated solution qf dinitroso-resorcin in 75 per cent, 

 alcohol ; then dehydrated, embedded, and sectioned. 

 A specific reaction for the neurokeratin network of 

 medullated nerves is obtained by this method. Rehm's 

 Method. For axis-cylinders. Stain sections for a few 

 minutes in a concentrated aqueous solution of Congo red, 

 wash in alcohol, treat for 10 minutes, until they become 

 blue, with acid alcohol, clear with origanum-oil, and 

 mount. This gives a clear axis-cylinder stain with con- 

 siderable other detail. Or, alcohol-hardened sections 

 may be stained for I or 2 days in a 0.5 per cent, aqueous 

 solution of hematoxylin, washed out in an aqueous solu- 

 tion of lithium carbonate until no more color is given off, 

 then dehydrated, and mounted. Counterstain for a few 

 minutes in a o.l per cent, aqueous solution of Bismarck 

 brown. Axis-cylinders, cells, and processes appear gray- 

 black. Sahli's Methods. 1. Sections of nerve-tissue 

 hardened in Midler's fluid are stained for a few minutes 

 or hours in a liquid prepared as follows : Mix 24 parts of 

 a saturated aqueous solution of methylene-blue, 16 parts 

 of a 5 per cent, solution of borax, and 40 parts of water ; 

 let the mixture stand a day, and filter. Wash sections 

 in water or alcohol until the gray matter can be distin- 

 guished from the white, clear in cedar-oil, mount in 

 balsam. Nerve-tubes and nuclei of neuroglia appear 

 blue, ganglion-cells greenish. Micrococci, if present, 

 are stained. 2. Sections hardened as detailed, and 

 washed for from 5 to 10 minutes in water, may be stained 

 for several hours in a concentrated aqueous solution of 

 methylene-blue. When they have acquired a deep-blue 

 color, rinse in water, and stain for 5 minutes in a satur- 

 ated, aqueous solution of acid fuchsin. Rinse in 

 alcohol, and differentiate in a liberal quantity of 

 water. The axis-cylinders appear red, the myelin- 

 sheaths blue. A still finer differentiation is obtained by 

 rinsing in alcohol containing from o.l to I per cent, 

 of a potassium-hydroxid solution, and then differen- 

 tiating in water. Clear with cedar-oil, mount in balsam 

 dissolved in cedar-oil. Schmaus' Method. For 

 axis-cylinders in the spinal cord. After hardening in 

 Midler's fluid, stain sections for from 15 to 20 minutes in 

 the following solution : sodium carminate I gm., uranium 

 nitrate 0.5 gm., water 100 c.c, heat for half an hour, 



and, when cold, filter. Wash out in water. Anothe: 

 stain that may be used is a o. 25 per cent, solution of 

 English blue-black in 50 per cent, alcohol, to which 1 

 little picric acid has been added. V. Thanhoffer'! 

 Methods. For multipolar nerve-cells. Press fresl 

 tissue between two cover-glasses, separate these, ant 

 let them dry in air. Float them on a concentrate! 

 aqueous solution of methylene-blue for several hours 

 Wash in water, pass through alcohol and clearing fluid 

 or dry in air. Mount in balsam (that is not dissolve! 

 in chloroform). Or, place fresh tissue for 3 or 4 days ii 

 Landois' fluid ; then stain in bulk for from 24 to 4 

 hours in equal parts of strong ammoniacal carmin an 

 methylene-blue solutions. Upson's Gold Methods 

 I. Harden the tissue in Midler's fluid for from 2 to 

 months ; then wash in water, and place for 2 days in 5 

 per cent, and then for 2 months in 95 per cent, alcohol 

 Embed in celloidin, and treat the sections with 80 pel 

 cent, alcohol for a few days before staining. The set 

 tion to be stained is first rinsed in water, then transferre 

 to a I per cent, aqueous solution of gold chlorid ft f 

 from 10 to 30 minutes, washed in water, immerse 

 for ]/% a minute in a 10 per cent, solution of sodiutj 

 hydroxid, again washed in water, and then place 

 in a reducing fluid consisting of 5 c - c - of sulphurot 

 acid, from 5 to IO drops of a 5 per cent, tincture 

 iodin, and 1 drop of a 37 per cent, solution of ferri 

 chlorid. When the sections assume a red color, the ( 

 should be removed from this fluid, and washed 

 dehydrated, and mounted in the usual way. 

 the sections, soon after cutting, in a 1 per cent, gold 

 chlorid solution for ^ an hour; wash in water, an 

 immerse for ^ a minute in a 15 per cent, solution 0] 

 sodium hydroxid, to which add, at the time of using, j 

 trace of chromic acid. Wash again, and place in a n 

 ducing fluid consisting of 15 drops of solution of stain: 

 chlorid, I or 2 gm., in 30 c.c. of a 1 per cent, tinctu 

 of iodin, distilled water 3 c.c, 3 drops of a 5 per ce: 

 solution of iron phosphate, and 3 c.c. of sulphurot 

 acid. 3. For axis-cylinders and nerve-cells. Hank 

 in the dark in a solution of potassium bichromate for 

 months, increasing the strength from I to 2.5 per cen 

 Wash and transfer to alcohol, increasing in strength fro , 

 50 to 95 per cent. Section, free or embedded, deh 

 drate, and put sections in a 1 per cent, gold-chlorid sol 

 tion with 2 per cent, of hydrochloric acid added 

 transfer on filter-paper to IO per cent, solution of pota-| 

 5 c.c, containing a trace of potassium feiricvani 

 After y z a minute, wash, and transfer to the follow ■ 

 ing : sulphurous acid 5 c.c; 3 per cent, tincture) 

 iodin from 10 to 15 drops; mix, and add solution' 

 ferric chlorid I drop. When the section has becot 

 colored, wash, dehydrate, clear, and mount. 4. >• ; 

 tions made as detailed are placed in a I per cen 

 chlorid solution 5 c.c. saturated solution of ammoniu 

 vanadate 10 drops, hydrochloric acid 3 drops, 

 after 2 hours' immersion, wash in distilled water, ai 

 place for from ^ to 1 minute in a mixture of 

 cent, caustic-potash solution 5 drops, IO per cent, poo 

 sium-permanganate solution 10 drops, and a trace 1 

 ammonium vanadate ; rinse in distilled water, and tre 

 until they become red with the following freshly pi 

 pared reducing mixture: 3 percent, tincture ol ic 

 to which has been added 15 drops of tin chlon 

 of distilled water, from 3 to 5 drops of a satun 

 tion of iron phosphate, and sulphurous acid 

 precipitate will be thrown down when those soltttio 

 are mixed, and at the instant that this occurs thi 

 should be put into it. The remaining treatment is M 

 the other methods. Van Gieson's Picro-acid-fuc 

 sin Method. Harden small pieces of nerve 

 Midler's fluid or alcohol, or both, and embed in celloidl 



