STAINS, ETC. 



1392 



STAINING OF NERVE-TISSUE 



through a spirit-flame, or place it for 2 minutes in 

 absolute alcohol. Treat with chloroform for 2 minutes, 

 rinse in water, and carry to 5 per cent, chromic acid 

 for 1 or 2 minutes, and wash in water. Pour a few 

 drops of carbol-fuchsin upon the preparation and heat 

 to boiling (which occurs in about a minute) ; drain and 

 decolorize in 5 per cent, sulphuric acid, wash thor- 

 oughly in water, and counterstain for half a minute with 

 methylene-blue or malachite-green in aqueous solution. 

 The spores are stained dark-red, the protoplasm of the 

 bacilli blue or green, according to the after-stain used. 

 3. Neisser's Method ( Sternberg ) . Pass the cover- 

 glass preparation three times through the flame of a 

 Bunsen burner, float upon a solution of anilin-fuchsin, 

 and heat to near the boiling-point for I hour ; wash in 

 water, and decolorize in hydrochloric acid 25 parts, 

 and alcohol 75 parts. Counterstain in a saturated aque- 

 ous solution of methylene-blue. The spores are stained 

 red, the protoplasm of the bacilli blue. Prolonged 

 action of the hydrochloric-acid solution will decolorize 

 the spores as well as the bacilli. 4. Place the slide or 

 cover-glass, with the culture dried upon it, in a hot- 

 air oven for 1 hour at 120 C, or for 15 minutes at 

 180 C, or the cover-glass may be passed 8 or 10 

 times through the flame of a Bunsen burner. Stain 

 in an aqueous solution of a basic anilin dye. The 

 spores alone are stained {Sternberg). Streptococcus 

 erysipelatosus (Fehleisen) may be stained with 

 the usual anilin dyes and by Gram's method. III. 

 Preliminary Methods. Biedert's Method. Used 

 in examining sputa which contain few tubercle-bacilli. 

 Mix 15 c.c. of the sputa with from 75 to loo c.c. of 

 water and a few drops of potassium or sodium hydroxid 

 solution. Boil until the sputa are thin. Place in 

 a conical glass vessel and after two days pour off the 

 supernatant liquid. Stain the precipitated sediment. 

 Kaatzer's Method. Mix the sputa with from a I to 

 a 3 per cent, solution of caustic soda or potash. This 

 disolves the cells and mucus, but preserves the elastic 

 fibers and bacteria. Stain the sediment. Clear the 

 preparation with a dilute solution of acetic acid. 

 Kiihne's Method. This method is used to over- 

 come the viscidity of sputum and to facilitate the 

 spreading of a thin and even film on the cover-glass. 

 It consists in adding to the sputa an equal volume 

 of a saturated solution of borax. A concentrated 

 aqueous solution of ammonium carbonate will reduce 

 the consistency of less viscid sputa. Muhlhausen's 

 Method. This method is used to render sputa less 

 viscid. It consists in adding to the sputum from 6 to 

 8 times its volume of a 2 per cent, solution of caustic 

 potash. Preserving Sputum. Savelieff' 's Method. 

 This is a process for preserving sputum for purposes 

 of subsequent examination. Let the patient expecto- 

 rate in a receptacle containing 95 per cent, alcohol, in 

 which the sputum may remain for several months, 

 and in which it is hardened by dehydration and 

 coagulation. A few drops of caustic-potash solution 

 added to a small lump of the hardened sputum on a 

 slide will liquefy it in a few minutes, and from this 

 the cover-glass preparations are made. When dry, 

 fix the film by passing the cover-glass thrice through 

 the flame of a spirit-lamp, wash in water to remove 

 the potash, and then stain according to any of the 

 given methods. Sectioning Sputum. Gabritschews- 

 k/s Method. Place the denser portions of freshly 

 expectorated sputum in Muller's fluid, or some other 

 hardening reagent, and then embed in celloidin. Stain 

 the sections in safranin, alum-carmin or hematoxylin- 

 eosin. Aronson and Philip treat the sputum first with 

 corrosive sublimate, and, according to Schmidt, it may 

 be embedded in paraffin as well as celloidin. 



STAINING OF NERVE-TISSUE. 

 Adamkiewicz's Method. Wash sections of spinal cord 

 in water, then in water acidulated with nitric acid, and 

 stain in a concentrated solution of safranin. Treat »vifh 

 alcohol and clove-oil until no more color is given off; 

 wash in water, then in, water acidulated with acetic 

 acid, stain in methylene-blue, and clear as before. This 

 process is said to demonstrate the " chromoleptic zones" 

 which surround the gray matter. The myelin ("ery- 

 throphilous substance " of Adamkiewicz) appears red, 

 the nuclei of nerves, neuroglia, and vessels appear violet. 

 This method is of value in the study of degenerative 

 changes, as the erythrophilous substance of pathologic 

 nerves does not take the stain. Alt's Method. 

 Adapted to the study of peripheral axis-cylinders. 

 Stain for two hours in a solution of Congo red in abso- 

 lute alcohol ; wash out in alcohol. Axis-cylinders 

 of, Centric Fibers (Bevan W. Lewis). Remove the 

 myelin from sections by prolonged immersion in water, 

 and then stain with anilin blue-black. The axis-cylin- 

 ders appear as slightly wavy, swollen bands. Ciaccio's 

 Method. This method is especially suitable for the ter- 

 minations of nerves in muscles and in the cornea. Place 

 small pieces of tissue, about 2 mm. cubes, for 5 min- 

 utes in the fresh, filtered juice of a lemon ; wash, and 

 place for from y 2 to I hour in a I per cent, solution of 

 gold and cadmium chlorids in the dark ; wash, and 

 carry to a I per cent, solution of formic acid for 24 hours 

 in the dark, then for 12 hours in sunlight; lastly, 

 for 24 hours in pure formic acid; wash, tease, 

 and mount in glycerin. Dausac's {A. Michel) 

 Method. Very minute pieces of tissue are fixed 

 in a watery solution of picric acid, chromic acid, and 

 nitric acid (the proportions are not given) for from X to 2 

 hours; washed in water for from ^ to 1 hour; em- 

 bedded in celloidin, cut, and placed in 90 per cent, al- 

 cohol. The sections are now transferred to Ehrlich's 

 fluid for from 2 to 5 minutes, rinsed in water, and placed 

 for from 2 to 5 minutes in a I per cent, solution of potas- 

 sio-gold chlorid ; rinsed in formic acid for I minute, car- 

 ried to a caustic-soda solution, I : 6, rinsed in water, and 

 placed in a IO per cent, solution of lithium carbonate tor 

 Yz hour. From this they are brought into a 10 per cent 

 solution of potassium iodid for from 2 to 3 minutes, and 

 are then reduced for IO minutes in a strong solution of 

 sodium thiosulphate. The axis-cylinders are stained a 

 black-violet; the remaining tissue is faintly col 

 Platinum chlorid or palladium chlorid gives the same 

 result. Dausac recommends this method also for fibrin 

 and elastic fibers. Exner's Method. A small piece 

 of the cortex cerebri, not exceeding 1 cubic cent; 

 in size, is placed in a relatively large volume ol 1 per 

 cent, osmic acid, which should be renewed every 2 day.-. 

 After from 5 to 10 days, wash with water, treat with alee 

 hoi, and embed. Treat sections on the slide with - 

 ammonia, which clears the tissue, and reveals the medul- 

 lated fibers stained black. Pieces of tissue, as fresh as 

 possible, not over )4 cm - m thickness, are placed in a 1 

 per cent, solution of osmic acid, the quantity of which 

 must be at least IO times the volume of the tissu 

 which should be renewed in 2 days. In 5 or 6 days 

 wash in water and embed. The sections are plai 

 the slide in glycerin to which a drop of ammonia has 

 been added (strong ammonia and water 1 : 5°)- 

 medullated fibers appear gray or black. The preparation 

 is not permanent. Free Nerve-endings in the Skin. 

 Place small cubes of the skin of the palmar surfa 

 the fingers or toes, all adipose tissue being removed, in 

 l>oiled gold chlorid and formic acid after this 111 

 has cooled. In an hour, transfer the tissues to slighl } 

 acidulated water and expose to sunlight until the 

 is reduced. Harden in alcohol, section, and mount in 



