Appendix B: Some Standard Reagents and Their Uses 181 



the particular element to be investigated (cell, axone, termina- 

 tion) has developed a well-marked blue color. It is important 

 to catch the color at the proper stage and fix it because it 

 soon begins to fade. 



Fixing the Stain. When the desired element has developed 

 a satisfactory blue color, the tissue is transferred immediately to 

 a saturated aqueous solution of Ammonium picrate (Dogiel's 

 method) and left for from 6 to 24 hours. For final mounting the 

 tissue should be teased out sufficiently to show the proper ele- 

 ments and then mounted in a few drops of a mixture of pure gly- 

 cerin (free from acid) and ammonium picrate (saturated aqueous 

 solution), equal parts. It is well to let the tissue stand in 20 to 

 30 volumes of this glycerin-picrate mixture for a day or two before 

 mounting it. If the preparation is to be kept the cover-glass 

 should be sealed (chap, xiii, II, A, 6). 



Sections. If it is desired to make paraffin sections and mount 

 them in balsam, after treatment with the ammonium picrate (10 

 to 15 minutes), the tissue must be placed into 20 or 30 volumes 

 of Bethe"s fluid, which renders the color insoluble in alcohol. 



Bethels fluid. 



Molybdate of ammonia 1 gram 



Chromic acid, 2 per cent, aqueous solution . 10 c.c. 



Hydrochloric acid, concentrated C. P. ... 1 drop 



Distilled water .... 10 c.c. 



The tissue is left in this mixture for from 45 to 60 minutes 

 (for small objects) and then washed 1 to 2 hours in distilled 

 water. Dehydrate directly in absolute alcohol ; follow this with 

 xylol, imbed in paraffin, and section in the ordinary manner. 

 Sections may be counterstained in alum-carmine or alum-cochineal. 



c) Immersion Method. Material which cannot be readily 

 injected or which has failed to stain may be stained by immersion. 

 A 0.1 per cent, solution of the stain is used (dilute 1 volume of 

 the solution used for injection with 9 volumes of normal saline). 

 To small pieces (2 to 3 mm. thick) of the tissue, add a few 

 drops of the stain at intervals of about three minutes. The tis- 

 sue should always be moist, but never covered sufficiently by the 

 solution to exclude air. Examine the preparation from time to 



