STAINING. 45 



To stain with this solution, one or two drops are placed on the slide 

 over the object and the whole put in a moist chamber for twenty-four 

 hours. A cover-slip is then placed over the preparation, the picrocarmin 

 drained off with a piece of blotting-paper, and a drop of formic-glycerin 

 (i : 100) brought under the cover-slip by irrigation. Proper differentia- 

 tion takes place only after a few days, and the acid-glycerin may then be 

 replaced by the pure glycerin. In objects fixed with osmic acid, the 

 nuclei appear red, connective tissue pink, elastic fibers canary yellow, 

 muscle tissue straw color, keratohyalin red, etc. 



Weigert's Picrocarmin. The preparation of Weigert's picro- 

 carmin is somewhat simpler. 2 gm. of carmin are stirred in 4 c.c. 

 of ammonia and allowed to remain standing in a well-corked bottle 

 for twenty-four hours. This is mixed with 200 c.c. of a concentrated 

 aqueous solution of picric acid to which a few drops of acetic acid are 

 added after another twenty-four hours. The result is a slight precipitate 

 that does not dissolve on stirring. Filter after twenty-four hours. Should 

 the precipitate also pass through the filter, a little ammonia is added to dis- 

 solve it. Both picrocarmin solutions dissolve off sections fixed to the slide 

 with albumen. 



Carmin=bleu de Lyon (of Rose). Sections or pieces of tis- 

 sue are first stained with carmin (alum- or borax-carmin). Bleu de Lyon 

 is dissolved in absolute alcohol and diluted with the latter until the solu- 

 tion is of a light bluish color. In this the sections or pieces of tissue are 

 after-stained for twenty-four hours (developing bone stains, for instance, 

 blue). 



Picric acid is often used as a secondary stain, either in aque- 

 ous (saturated solution diluted i to 3 times in water) or in alco- 

 holic solution (weak solutions in 70%, 80%, and absolute alcohol). 

 Sections previously treated with carmin or hematoxylin are stained for 

 two to five minutes, washed in water or alcohol, and transferred to abso- 

 lute alcohol, etc. Sections stained in safranin can be exposed to the ac- 

 tion of an alcoholic picric acid solution. A solution of picric acid in 

 70% alcohol may be used to wash sections stained in borax-carmin. 

 This often gives a good double stain. Sections can also be first treated 

 with picric acid and afterward stained with alum-carmin. 



Hematoxylin. Van Gieson's Acid fuchsin-picric acid Solu- 

 tion. Stain in any one of the hematoxylin solutions and after rinsing 

 sections in water counter-stain in the following : 



Acid fuchsin, \% aqueous solution .... 5 c.c. 

 Picric acid, saturated aqueous solution . . loo " 



Dilute with an equal quantity of distilled water before using. The 

 hematoxylin stained sections remain in the solution for from one to two 

 minutes, are then rinsed in water, dehydrated and cleared. 



Hematoxylin=eosin. Sections already stained in hematoxylin 

 are placed for two to five minutes in a i % to 2 % aqueous solution of 

 eosin or in a i% solution of eosin in 60% alcohol. They are then 

 washed in water until no more stain comes away, after which they remain 

 for only a short time in absolute alcohol. In place of the eosin solution 

 a i % aqueous solution of benzopurpurin may be used or the following 

 solution of erythrosin (Held) : 



Erythrosin I gm. 



Distilled water 150 c.c. 



Glacial acetic acid 3 drops. 



