PREPARATION OF MEDIA 1I44-II 



ferred to complete the sterilization in one day, heat for 15 minutes at 15 pounds without 

 letting the air escape; then let the air escape slowly while maintaining pressure, after 

 which the air vent should be closed and sterilization continued at 15 pounds for 16 

 minutes longer. After completion of the sterilization the pressure should be allowed to 

 fall very slowly. 



Use: Cultivation of diphtheria organism. 



Hunloons Hormone Heart Infusion Broth {L&S No. S'37). Mix 10 g. Bacto peptone, 

 10 g. gelatin, 5 g. NaCl, one whole egg and 500 g. of finely chopped beef heart in a 

 litre of water. Place in an enamel-ware vessel, e.g. a large coffee pot. Heat over a free 

 flame with constant stirring until the red color of the meat infusion changes to brown at 

 a temperature of about 68°C. Do not go beyond this temperature. Adjust to slightly 

 alkaline to litmus and then add 1.0 ml. additional A^/1 NaOH per litre of medium. 

 Cover the vessel and place In an Arnold sterilizer or in a water bath at 100° for one 

 hour. Remove the vessel from the sterilizer and separate with a glass rod the firm clot 

 which has formed from the side of the vessel. Return to the Arnold sterilizer at 100° 

 for 11^4 hours. Remove the vessel and allow to stand at room temperature for about 

 10 minutes in a slightly' inclined position. Pipette ofif the fluid portion or decant. If it 

 is poured through a fine wire sieve, many of the fine pieces of meat clot may be caught. 

 (Avoid filtering through cheese cloth, cotton or other absorbent materials.) Allow it to 

 stand in tall cylinders for 15 to 20 minutes until the fat present has risen to the surface 

 and been removed. The medium may be further cleared by filtering through glass 

 wool, asbestos wool, sedimentation or centrifugation. Add 0.15% dextrose and enough 

 laked blood to give a slight pink tint. Tube in 10 ml. lots. Sterilize by the inter- 

 mittent method. 



Use: To cultivate highly pathogenic organisms. 



Hitchens' Semi-solid Glucose Agar {Mulsow) (L &S No. 879) . Add 500 ml. of water to 1 

 pound of ground lean beef, and allow to stand at 37°C. for 48 hours. Express the juice 

 and add 20 g. peptone, 2 g. KNO3 and an equal amount (500 ml.) of a 0.2% agar so- 

 lution heated and cooled to 60°C. Adjust the reaction to + 0.9 to phenolphthalein. 

 Heat in the autoclave at 15 pounds pressure for 25 minutes. Filter and readjust the re- 

 action if necessary. Add 2 g. glucose. Final method of sterilization not specified. 



Use: Cultivation of gonococcus and microaerophilic bacteria in general. 



Egg Medium vnth Glycerol. Break several eggs into a graduated cylinder. Add y^ 

 as much meat infusion or Douglas' broth as the amount of eggs. Add 1% glycerin. 

 Stir to mix, taking care not to cause the formation of air bubbles or frothing. Filter 

 through gauze. Tube and slant in a pan or rack. Sterilize in the autoclave at 15 

 pounds for 15 minutes. 



Use: Cultivation of tubercle organism. 



2. DIFFERENTIAL MEDIA 



Endo medium. Dissolve 5 g. beef extract and 10 g. peptone in 

 1000 ml. water. Add 30 g. agar and cook in autoclave 45 minutes at 

 15 lbs. pressure. Filter; then add 10 g. lactose and sterilize 15 

 minutes at 10 lbs. pressure in small containers 100ml. in each. Just 

 before use prepare a 3% solution of basic fuchsin (85-90% actual dye 



