DECALCIFYING FLUIDS. 405 



It is a solution of 30 grams of sodium sulphate and 60 grams of potassium 

 bichromate in 3000 cc. of water. To make Orth's fluid, 10 cc. of formaline 

 are added to 100 cc. of Miiller's fluid shortly before using. Small blocks 

 of tissue should remain in it for 3 or 4 days, when, after washing thoroughly 

 in running water, they are put in 80 per cent, alcohol. 



Alcohol. The higher grades of alcohol are important fixing fluids, 

 although for most purposes inferior to Zenker's fluid or formaline. Tissue 

 may be put directly into 95 per cent, or absolute alcohol, a piece of ab- 

 sorbent cotton being under it. The alcohol should be changed after 3 or 

 4 hours, and after 3 or 4 days the tissue is transferred to 80 per cent. 

 Some histologists recommend passing the fresh tissue through graded alco- 

 hols before putting it in absolute; this causes less shrinkage but is said to 

 fix imperfectly. One may begin with 80 per cent. 



Specimens should be kept in 80 per cent, or 90 per cent, alcohol after 

 they have been preserved. They macerate in the weaker alcohols and lose 

 their staining capacity in those which are stronger. 



D ECALCIFICATION . 



Specimens which contain bone or calcareous material cannot be 

 sectioned until they have been decalcified, which can be done only after 

 they have been fixed, and hardened for a few days in alcohol. They are 

 then placed in considerable amounts of dilute nitric acid (3 to 5 cc. of con- 

 centrated nitric acid in 100 cc. of water). This should be renewed for 3 or 

 4 days, until the bone can be cut with a scalpel or be penetrated easily with 

 a needle. The acid is removed from the tissue by immersion in running 

 water for a day, and the block is returned to the alcohol. 



Phloroglucin is sometimes added to the decalcifying fluid to protect the 

 tissue, and the nitric acid may be diluted with alcohol. The following 

 solution has been recommended: 



Phloroglucin i 



Nitric acid 5 



Alcohol 70 



Water 30 



A slight addition (i or 2 per cent.) of nitric or hydrochloric acid to 

 80 per cent, alcohol may be used in decalcifying small embryos. 



IMBEDDING. 



Imbedding is the process by which blocks of fixed, hardened, and 

 decalcified tissues are prepared for sectioning. Sometimes the tissue is 

 stained before being imbedded, as will be described later; often all the 

 staining is done after the sections have been cut. Imbedding consists in 



