STAINS, ETC. 



1395 



STAINING OF NERVE-TISSUE 



e ; then bring them for from 5 to 10 minutes into I 

 jart of a saturated aqueous solution of lithium-carbonate 

 ind 3 parts of water. Stain for from 18 to 24 hours in 

 Japanese red-wood solution (see formula in Flechsig's 

 method), and differentiate in Weigert's decolorizing mix- 

 FlescK s Modification. Celloidin or other sections 

 Sire put for a few minutes or more in a 0.5 percent, 

 htomic-acid solution, then rinsed in water, and brought 

 ' ato the jtain. Decolorize in the usual way-. This method 

 las the advantage of staining more rapidly and producing 

 utter differentiation of the nerve-cells, especially in the 

 i leripheral ganglia, and also of giving differentiation of 

 he medulla of central and peripheral nerves. 5. 

 Modification. Small segments of fresh cord 

 ,.re put for 2 days into a saturated, aqueous solution of 

 ' leutral copper acetate, then for from a day to a day and a 

 lalf into a 5 per cent, or a saturated solution of potassium 

 lichromate; rinsed in water, and placed in 70 per cent, 

 i Jcohol, for from 36 to 48 hours, in the dark ; then treated 

 jor the same period with absolute alcohol, in the dark, 

 md embedded. Treat paraffin sections with alcohol, then 

 vater, and stain for from 15 to 30 minutes in a well- 

 ipened mixture of I part hematoxylin, I partammonium- 

 .lum, 30 parts alcohol, 300 parts water. Rinse in water, 

 lifferentiate in acid alcohol until a red color appears, wash 

 vith water until they turn blue or bluish-gray, counter- 

 tain, if desired, by momentary immersion in a neutral 

 armin solution, and mount. 6. Hill's Modification. 

 : 'ieces of nervous tissue are put in from a 2 to a 2.5 per 

 ent. solution of potassium bichromate for 6 weeks ; then 

 vashed daily in 30 per cent, alcohol until the fluid re- 

 nains clear ; then fully hardened in strong alcohol. Wash 

 n water small pieces and put them in solution of car- 

 nin and alum prepared thus : Boil for 3 hours, in water 

 n which carmin and potash-alum have been placed, 

 md restore the water lost by evaporation. Both car- 

 nin an 1 alum should be in excess. Filter when cold. 

 j Vfter 2 days put the pieces for 24 hours in a half-satu- 

 ated solution of copper acetate, then into hematoxylin 

 or 8 hours at 40 C. Decolorize in Weigert's decol- 

 irizing fluid. The nerve-cells and non-medullated 

 ibers are rendered susceptible of staining by the alum- 

 armin. 7. Kultschi'zky's Modification. Harden for 

 me or two months in Erlicki's solution, embed in celloi- 

 lin and photoxylin, and cut. Stain sections for from 1 



24 hours in hematoxylin 1 gm., dissolved in a little 

 Icohol and added to 100 c.c. of 2 per cent, acetic acid. 

 ■Vash out in a saturated solution of sodium or lithium car- 

 ■onate. A finer differentiation is obtained by decoloriz- 

 ng in a lithium-carbonate solution containing 10 per cent, 

 if potassium ferricyanid. 8. Lissauers Modification. 

 V. rapid method which gives good results with cerebral 

 issue even when it is imperfectly hardened. Place 

 ections that have been hardened in Miiller's fluid in a 

 - per cent, solution of chromic acid, and heat until 

 •ubbles begin to form ; rinse in water, place in Weig- 

 •rt's hematoxylin, and heat again till bubbles begin to 

 orm. Differentiate by Pal's method. 9. PaT s Mod- 

 fication. After staining in the hematoxylin solution, 

 he sections are washed in water, to which, if they are 

 iot stained a deep-blue, a trace of lithium carbonate is 

 dded. They are next placed in a 0. 25 per cent, solu- 

 ion of potassium permanganate for half a minute, 

 >nsed in water, and then brought into a decolorizing 



1 olution composed of 1 part each of chemically pure 



:alic acid and potassium sulphite, and 200 parts of 



'istilled water. In a few seconds the gray substance is 



rolorized, the white remaining blue. Wash well in 



iter, and double-stain with eosin or picrocarmin. 10. 



Modification. Harden in a solution of I gm. of 



nromic acid and 5 gm. of copper acetate in loo c.c. 



»' water. Dehydrate, and embed in celloidin. Stain 



for 2 hours in hematoxylin (7 or 8 drops of a 5 per cent., 

 alcoholic solution to 30 c.c. of alcohol). Differentiate in 

 acid alcohol, wash out for 20 minutes in water, dehy- 

 drate, and mount. Double-stain if desired in borax-car- 

 min. 11. Schafers Modification. Harden the tissue for 

 from 4 to 6 weeks, and put the sections in Marchi's fluid 

 ( 1 part of a I per cent, osmic-acid and 2 parts of a 3 per 

 cent, potassium-bichromate solution). Wash quickly in 

 water, and stain in hematoxylin I gm., acetic acid 2 c.c, 

 water 100 c.c. Differentiate by Pal's method. This 

 method is applicable when sections have been too long in 

 alcohol. 12. VasaWs Modification. From alcohol the • 

 sections are transferred to a solution of hematoxylin I gm. 

 to loo c.c. of water, and decolorized by the aid of heat. 

 After from 3 to 5 minutes they are put into a saturated, 

 filtered solution of copper acetate and left for the same 

 length of time, when they become black. They are washed 

 in water and placed in a solution of borax 2 parts, potas- 

 sium ferricyanid 2.5 parts, and water 300 parts, in which 

 the degenerated areas, the cells, and the neuroglia be- 

 come decolorized, the medullated fibers remaining dark. 

 After decolorization, wash in water, dehydrate, clear, 

 and mount. Counterstaining by picro-carmin or alum- 

 carmin may be practised. 1 3. Wolter s Modification. 

 I. Stain sections in a solution of 2 gm. of hematoxylin 

 in a little alcohol and 100 c.c. of 2 per cent, acetic acid 

 at 45 C. for 24 hours. Dip them in Midler's fluid, and 

 differentiate by Pal's method. This is an intense myelin 

 stain ; medullated fibers appear blue-black, ganglion-cells 

 yellow, the ground light. Or, sections of tissue hardened 

 in Miiller's fluid and cut in celloidin are mordanted for 

 24 hours in a mixture of 2 parts of 10 per cent, vanadium 

 chlorid and 8 parts of 8 per cent, aluminum acetate ; 

 then washed for 5 or 10 minutes in water, and stained 

 in the foregoing solution of hematoxylin and differenti- 

 ated with Weigert's decolorizing fluid. This is a myelin 

 stain, with a splendid differentiation of the processes 

 of Purkinje's cells. Monti's Copper Method. Small 

 pieces of nervous tissue are hardened in a 2 or 3 per cent, 

 solution of potassium bichromate or Miiller's fluid until 

 they are quite firm. They are then immersed in a mix- 

 ture of equal parts of copper sulphate and Miiller's fluid. 

 A reaction takes place which stains the nerve-cells a red- 

 dish color in direct, or a blackish-yellow color in trans- 

 mitted light. Nerves and Nerve-cells in a Frog's 

 Heart. Find the sinus venosus, and ligate the inferior 

 and two superior vena; cava; opening into it ; make an 

 incision into one of the aortae, and into it tie a fine, 

 glass cannula. Inject normal saline solution to wash out 

 the cavities of the heart. Distend the cavities with 2 

 per cent, gold chlorid 4 parts and formic acid I part, pre- 

 viously boiled together and cooled. Ligate the other 

 aorta, adjust a ligature below the cannula, cut out the 

 heart, and place it for from a ^ to I hour in 5 c.c. of the 

 gold mixture. Open the auricles, wash the heart in water, 

 and expose it to light in distilled water 50 c.c, con- 

 taining 3 drops of acetic acid. Reduction of the gold 

 takes place in from 3 to 4 days. Examine the auricular 

 septum in glycerin, for pyriform nerve-cells with straight 

 and spiral nerve-processes. A 2 per cent, solution of 

 osmic acid, used in place of the gold solution , brings the 

 nerve-fibers into prominence. Nigrosin Method. For 

 axis-cylinders. Stain sections for from 5 to 10 minutes 

 in concentrated aqueous solution of nigrosin, decolorize 

 in dilute, then in absolute alcohol, and clear in origanum- 

 oil. Nikiforoff's Modification of Adamkiewicz's 

 Method. Harden in a chrome-salt and transfer directly 

 to alcohol. Section, and place in alcohol. From the al- 

 cohol sections are brought for 24 hours into a concentrated 

 aqueous solution of safranin, or anilin-water safranin, or 

 in5percent. carbolic acid and safranin. Differentiate in 

 alcohol until the gray is distinguishable from the white 



