252 MICROBIOLOGY 



meat (veal or beef) and not meat extract. The bouillon shall 

 contain 1% peptone ("Witte's), 0.5% sodium chloride or acid 

 potassium phosphate, chemically pure glycerin 5% (this may 

 vary from 3 to 7%). The final reaction of the bouillon shall 

 be 0.75 to 1.0 to phenolphthalein, the normal acidity of the 

 broth being corrected by the addition of sodium hydrate solu- 

 tion. 



2. ' ' The culture of tubercle bacillus employed shall be of 

 mammalian origin and shall produce an active tuberculin. 



3. "The containers in which the cultures of tubercle 

 bacilli are grown shall be in form similar to the Erlenmeyer 

 flasks of the Jena pattern, preferably 300 cc. capacity, each 

 flask to contain 100 cc. of bouillon. 



4. ' ' The cultures should be allowed to grow for four weeks 

 after the surface of the medium is completely covered with the 

 growth of the tubercle bacillus. 



5. "In the preparation of the tuberculin the ripe cultures 

 are shaken down, placed in a steam sterilizer and subjected to 

 live steam for at least 2% hours, filtered through paper, 

 evaporated to one-tenth of original volume. Before being 

 supplied to practitioners the tuberculin should be passed 

 through Chamberland or Berkefeld filters. 



6. * ' Tuberculin should be sent out from the laboratory so 

 that the practitioner need not dilute it before using. The dilu- 

 tions when exposed to the air should not for human practice be 

 prepared for any great length of time before use. The dilu- 

 ent suggested is 0.25% phenol in physiological normal sodium 

 chloride solution." 



New tuberculin. In 1897' Koch 18 described a new tu- 

 berculin, prepared by grinding in a mortar tubercle bacteria 

 that had been dried in vacuo (this is done by machinery where 

 large quantities are made). The grinding is continued until 

 the microscopic examination fails to show unmutilated bac- 

 teria. One gram of the dry mass is shaken up in 100 cc. of 

 sterile distilled water. This mixture is then centrifugalized at 



18 Koch. Deut. med. Woch., Bd. XIV (1897). 



