HYPERSENSITIVENESS 507 



determined whether for routine preparations this additional 

 washing process is absolutely essential before precipitation. In 

 order to obtain the quantitative results given in this paper, it 

 was, of course, necessary.) The protein is then precipitated from 

 the ])ure colloidal solution in a final concentration of 10 per cent 

 trichloracetic acid until the wash is clear and colorless. The 

 drained precipitate is partly dried in vacuo and then ground to 

 a fine powder under larger volumes of ether, which simultaneously 

 dehydrates the product and removes the trichloracetic acid. A 

 fine cream-colored powder, TPT, is thus quickly obtained." 



The product is rendered completely soluble in water by cau- 

 tiously adding a few drops of N/10 alkali and immediately neu- 

 tralizing it with HCl. They prepare stock solutions in 0.9 per 

 cent saline containing 0.5 per cent phenol. 



The underlying principles of the above technique may be sum- 

 marized as follows : 



1. Using a standard virulent culture of the tubercle bacillus and 

 a synthetic medium, a tuberculin is produced. 



2. The bacteria are removed by filtration. 



3. By means of ultrafiltration the resulting tuberculin is con- 

 centrated and at the same time rendered free of all substances 

 except the active colloidal tuberculin protein and the ever present 

 polysaccharide found in old tuberculin. 



4. The active tuberculin protein is separated from the poly- 

 saccharide by precipitation with trichloracetic acid. 



5. The precipitate is washed, dried, and dissolved in saline to 

 which phenol is added as a preservative. 



The above treatment does not denature or in any way injure 

 the tuberculin protein. 



Seibert's sot and SOTT or PPD.— Seibert prepared also an 

 old tuberculin (SOT) from cultures grown in a synthetic medium. 

 She next concentrated and purified SOT by ultrafiltration and 

 precipitated the active principle with trichloracetic acid. The pre- 

 cipitate was treated with ether to remove the acid and to dehydrate 

 the protein. This purified protein derivative of the tubercle 

 bacillus she calls SOTT (synthetic medium old tuberculin pre- 

 cipitated with trichloracetic acid). In 1932 she prepared SOTT 

 from cultures grown on Long's medium, but later she has adopted 

 the .synthetic medium used by the Bureau of Animal Industrv. 



