SECTION TWO 



between bone and cartilage and for demonstrating the relative 

 amount of ossification. 



The removal of the viscera is unnecessary in the case of museum 

 specimens. 



Solutions required: 



A. Toluidine Blue . . . . . . 0-25 gm. 



Alcohol 70% . . . . . . 100 ml. 



Hydrochloric acid 0-5% . . 2 ml. 



Allow the solution to stand for twenty-four hours ; 

 then filter and store in a tightly corked bottle. 



B. Potass, hydroxide 2% aqueous . . 



C. Alizarin Red, S . . . . . . o-ooi gm. 



Potass hydroxide 2% aqueous . . 100 ml. 



(This solution should be freshly prepared.) 



D. Methyl salicylate 25% in cellosolve 



E. Methyl salicylate 50% in cellosolve 



F. Methyl salicylate 75% in cellosolve 



Technique: 



1. Wash specimens for twenty-four hours in 70% alcohol con- 

 taining 0-2% of concentrated ammonia solution „ 



2. Stain for seven days in Solution A. 



3. Harden and destain for seventy- two hours in four changes of 

 95% alcohol. 



4. Macerate for five to seven days, depending on the size of the 

 animal, in several changes of 2% aqueous potass, hydroxide. 

 {Note: This process is hastened by exposure to sunlight.) 



5. Transfer to Solution C for about twenty-four hours when the 

 bones should be well stained. If the specimen has been insuffi- 

 ciently macerated the soft tissue will be slightly stained, in which 

 case the specimen may be destained rapidly in acid alcohol (1% 

 sulphuric acid in 95% alcohol). 



6. Dehydrate by leaving the specimen in three changes of 

 cellosolve for six hours in each. Instead of cellosolve, 50%, 80% 

 and 90% alcohol, followed by three changes of benzol may be 

 used for dehydration. Small embryos require less time in the 

 dehydrating fluids. 



^ 55 



