THE PREPARATION OF CULTURE MEDIA 16 



of the organism in question. The substances most commonly used 

 for such enrichment are glucose, nutrose (sodium casemate), gly- 

 cerin, sodium formate, and uiisolidified animal proteins. As animal 

 and blood serum and whole blood must frequently be used in this 

 way, an understanding of the methods employed in obtaining these 

 substances is necessary. 



Method of Obtaining Blood and Blood Media. Blood serum from 

 beef and sheep may be collected in the manner recommended for 

 the collection of such serum in the preparation of Loeffler's medium, 

 pipetted into test tubes, and sterilized in the fluid state by exposure 

 to temperatures ranging from 60 to 65 C., for one hour upon six 

 consecutive days. It is not a simple matter to sterilize serum in 

 this way and requires much time and care. 



The method most commonly employed, in laboratories which have 

 access to hospitals, for obtaining clear serum depends upon the col- 

 lection of exudate or transudate fluids by sterile methods directly 

 from the pleural cavity, the abdominal cavity, or the hydrocele 

 cavity. Sterile flasks or test tubes are prepared and the fluid is 

 allowed to flow directly out of the cannula into these. It is necessary 

 to avoid carbolic acid or other disinfectants in sterilizing instru- 

 ments and rubber tubing used during the operation. These should 

 be brought into the ward in the water in which they have been 

 boiled and not in strong antiseptic solutions, as is frequently done. 

 The fluid so obtained may be incubated and the contaminated tubes 

 discarded. The serum may then be added, in proportions of one to 

 three, to sterile broth or melted agar. 



Agar thus used is melted and cooled to 60 C., or below. One- 

 third of its volume of warmed exudate fluid is added, and the plates 

 are poured. 



Serum may be rendered free of bacteria by filtration through a 

 Berkefeld or Pasteur-Chamberland filter. This is an effectual 

 method, but requires much time and care. 



Whole blood may be obtained for cultural purposes by bleeding 

 rabbits or dogs or other animals directly from a blood-vessel into 

 tubes of melted agar. In the case of a rabbit, after the administra- 

 tion of an anesthetic (ether), an incision is made directly, over the 

 trachea, and, by careful section, the carotid artery is isolated, lying 

 close to the side of the trachea. Blood may be collected and 

 hemolysed by the gradual addition of the smallest amount of ether 

 which will completely hemolyse the amount treated. This may be 



