METhIdDS of staining. 367 



8. Dissolve off the paraffin in turpentine. 



9. Put in absolute alcohol for three to five minutes. 



10. Clear in turpentine or chloroform. 



11. Mount in dammar varnish. (Canada balsam is undesirable, because it 

 becomes much discolored with age.) 



Methods of Staining. 



In embryological work the specimens are usually stained in toto before im- 

 bedding, either with alum cochineal or with borax carmine, the former being the 

 more generally useful stain. Staining on the slide is also much used either to 

 secure a counterstain after the in toto coloration or to secure some special result. 

 For counterstains eosin, Lyons blue, and orange G are particularly recom- 

 mended. A few of the most important special stains are given below. 



1. Alum Cochineal. 



Formula : Powdered cochineal, 6 gm. 



Potassic alum, 6 gm. 



Water, . 80 c.c. 



Boil vigorously for twenty minutes; allow the fluid to settle; decant the clear 

 fluid; add more water and boil again; filter the entire solution and evaporate 

 the filtrate to 80 c.c; add a small piece of thymol or camphor to prevent the 

 growth of fungi. Alum cochineal is, on the whole, the best reagent for in toto 

 staining, as it will penetrate quite large objects and color them uniformly 

 throughout, and gives a good differentiation of the tissues. 



For in toto staining place the specimen in water until it sinks ; then transfer 

 it to the cochineal for twenty-four hours, or for large specimens longer; the 

 depth of the stain will depend upon the strength of the solution; transfer to 

 clean water for fifteen to twenty minutes to extract the alum, which otherwise 

 will crystallize in the tissues when the specimen is placed in alcohol ; the object 

 must not be left too long in water, because it extracts the color also ; put in 50 

 per cent, alcohol for one hour, then successively in 70 per cent., 80 per cent., and 

 95 per cent., when the specimen will be ready for imbedding. 



2. Borax Carmine. 



Forviula : Best carmine, 3 gm. 



Borax, 2 gm. 



Water, 50 c.c. 



Boil for twenty minutes ; allow the solution to cool ; add water enough to restore 

 that lost by evaporation, then add 50 c.c. of 70 per cent, alcohol, let the solution 



