CULTURE MEDIA FOR CULTIVATION OF MICRO ORGANSIMS 



579 



Preparation : 



(1) Dissolve 2, 3, and 4 in 1. 



(2) Add 1.5 g. trypsin and make slightly 

 alkaline. 



(3) Add chloroform. 



(4) Incubate at 37°C. for 48 hours and 

 then boil several minutes to remove 

 the chloroform. 



(5) Add 15.0 g. agar. 



(6) Adjust the reaction to pH 7.0 and add 

 15.0 g. lactose and 50.0 cc. of a 2.0% 

 sodium oleate solution. (In another 

 part of the article the author states 

 that best growth was obtained with 

 about 1.0% sodium oleate. The 

 percentage used above is only 0.1%. 



(7) Filter. 



(8) Tube. 



Sterilization: Sterilize in the autoclave. 

 Use: Cultivation of Lactobacillus hulgari- 



cus and Lactobacillus acidophilus. 

 Reference: Hunter (1924 p. 3). 



1852. Zoller's Citrate Milk Agar 



Constituents: 



1. Distilled water 5000.0 cc. 



2. Bacto Nutrient Agar (1.0%) 120.0 g. 



3. Sodium citrate 2.0 g. 



4. Milk (skimmed) powder. . . . 25.0 g. 



5. Na2HPOr2H20 6.0 g. 



6. Lactose 5.0 g. 



Preparation : 



(1) Stir 120.0 g. of Bacto nutrient agar 

 (1.0%) into 4 liters of distilled water. 



(2) Add 2.0 g. sodium citrate and set in 

 streaming steam in the autoclave 

 for about 5 minutes. 



(3) Stir 25.0 g. of any good grade of 

 milk powder in 100.0 cc. distilled 

 water. 



(4) Stir 6.0 g. Na2HP04-2H20 in 30.0 

 cc. of distilled water and heat to 

 boiling. 



(5) Add (4) to (3) and steam for 5 

 minutes at the same time the agar is 

 being steamed. 



(6) Remove the agar from the autoclave 

 and stir. 



(7) Add 50.0 cc. distilled water to (5) 

 following steaming. 



(8) Replace (6) and (7) in the autoclave 

 and heat for 5 minutes at 5 pounds 

 pressure. 



(9) Filter the milk phosphate solution 

 thru a small piece of absorbent 

 cotton placed in the neck of a funnel. 



(10) Add 5.0 g. lactose to filtered (9) and 

 make up to one liter by the addition 

 of distilled water. 



(11) Mix (10) and the agar from (8). 



(12) Distribute as desired. 

 Sterilization: Autoclave at 15 pounds pres- 

 sure for 15 minutes, or sterilize frac- 

 tionally by heating at 5 pounds pressure 

 for 10 minutes on each of 2 successive days 

 holding the medium at 30°C. between 

 heating. 



Use: Qualitative milk counts. 

 Reference: Zoller (1923 p. 385). 



1853. Smillie's Tissue Infusion Agar 



Constituents : 



1. 2.0% veal infusion agar 100.0 cc. 



2. Ascitic fluid 200.0 cc. 



3. Rabbit tissue (kidney) 

 Preparation : 



(1) Prepare 2.0% veal infusion agar. 



(2) Adjust (1) to 4-0.5 to phenolphthalein 



(3) When ready for use, melt sterile (2), 

 boil and cool rapidly to 50°C. 



(4) Warm, clear, straw colored, bile 

 free, ascitic fluid having a specific 

 gravity of 1.015 or higher to 45°C. 



(5) Place in a vacuum to remove air. 



(6) Mix one part melted (3) at 50°C. and 

 two parts (5) at 45°C. 



(7) Add 15.0 cc. of (6) to a sterile tube 

 containing sterile rabbit tissue 

 (kidney) (and the poliomyelitis 

 materials). 



Sterilization: Method not specified. 



Use: Cultivation of globoid bodies of 

 poliomyelitis. Author reported that the 

 organisms did not attack any added 

 carbohydrate. Other investigators used 

 similar media for the cultivation of 

 anaerobes. 



Variants : 



(a) Olitsky and Gates cultivated ana- 

 erobic organisms found in influenza 

 on a medium prepared as follows: 



(1) Prepare a 2.0% beef infusion agar. 



(2) Adjust (1) to pH = 7.4. 



(3) Mix one part (2) with two parts 

 sterile human ascitic fluid in a 

 flask at 40°C. 



