METHODS OF IMPREGNATION. 51 



fresh, the specimens must not exceed 3 or 4 mm. in thickness, and for 

 every piece of tissue treated about 10 c.c. of the osmium-potassium 

 bichromate mixture should be employed, the specimens remaining in the 

 latter (in the dark) at a temperature of 25 C. for a length of time vary- 

 ing according to the result desired (two or three days for the neurogliar 

 cells, from three to five days for the ganglion cells, and from five to seven 

 days for the nerve-fibers of the spinal cord). The objects are now dried 

 with blotting-paper or washed quickly in distilled water and then placed 

 for two or three days in a 0.75% silver nitrate solution at room-tempera- 

 ture. In this they may remain for four or five days without damage, but 

 not longer, as otherwise the precipitate becomes markedly granular (vid. 

 v. Lenhossek, 92). 



If Golgi's method be unsuccessful (this applies to all its modifica- 

 tions), the preparations may be transferred from the silver nitrate solu- 

 tion back into a potassium bichromate-osmic acid mixture containing 

 less osmic acid, in which they remain several days, and are then again 

 placed in the silver nitrate solution for from twenty-four to forty-eight 

 hours. This procedure may even be repeated. 



Cox obtains a precipitate in both cells and fibers by treating 

 small pieces of the central nervous organs with a mixture composed of 

 potassium bichromate 20 parts, 5 % corrosive sublimate 20 parts, distilled 

 water 30 to 40 parts, and 5% potassium chromate of strong alkaline 

 reaction 16 parts. The specimens remain in this mixture from one to 

 three months, according to the temperature, and are then further treated 

 according to Golgi's method. 



As the chrome-silver preparations are not permanent, and can not, 

 therefore, be subsequently stained, Kallius has suggested that the chrome- 

 silver precipitate be reduced to metallic silver by treatment with the 

 "quintuple hydroquinon developer" (hydroquinon 5 gm., sodium 

 sulphite 40 gm., potassium carbonate 75 gm., and distilled water 250 

 gm.). For this purpose 20 c.c. of the solution are diluted with 230 c.c. 

 of distilled water ; this mixture may be preserved in the dark for some 

 time if desired. Before using this latter solution, it should be mixed with 

 y$, or at the most y^, of its volume of absolute alcohol. The sections are 

 placed in a watch-crystal containing some of the latter mixture until they 

 turn black (a few minutes). As soon as the silver salt is completely 

 reduced, the sections are placed for from ten to fifteen minutes in 70 % 

 alcohol, then for five minutes in a 2o r / c solution of sodium hyposulphite 

 and, finally, washed for some time in distilled water, after which they 

 may be stained, and even treated with acid alcohol and potassium 

 hydrate. 



The following simple method for permanently mounting Golgi prepar- 

 ations under a cover-glass has been recommended by Huber. 



After impregnation with chrome-silver the tissues are hastily dehy- 

 drated, imbedded in celloidin, and cut in sections varying from 25 ft to 

 100 fi in thickness. The sections are then dehydrated and placed for 

 from ten to fifteen minutes in creosote, from which they are carried 

 into xylol, where they remain another ten minutes. The sections are 

 then removed to the slide. The xylol is then removed by pressing sev- 

 eral layers of filter-paper over the section. On removing the filter-paper 

 the sections are quickly covered by a large drop of xylol balsam and the 

 slide is carefully heated over a flame for from three to five minutes. Be- 



