704 



CULTURE MEDIA FOR CULTIVATION OF MICROORGANISMS 



4. Nutrose (10.0% solution) 



5. Malachite Green 

 Preparation : 



(1) Prepare a beef infusion of 1 pound in 

 2 liters water. 



(2) Add 30.0 g. agar. 



(3) Add 7.5 CO. N/1 HCl. 



(4) Boil 30 minutes. 



(5) Add 7.0 cc. N/1 NaOH. 



(6) Neutralize to litmus with sodium 

 carbonate. 



(7) Add 5.0 cc. N/1 sodium carbonate. 



(8) Add nutrose to make 1.0% solution 

 using a 10.0% solution of nutrose for 

 this purpose. (0.5% peptone may 

 be used instead of nutrose.) 



(9) Boil. 



(10) Distribute in 500.0 cc. flasks. 



(11) Let cool slowly in sterilizer following 

 sterilization. 



(12) Decant clear supernatant liquid. 



(13) To each 100.0 cc. agar add 2.0 to 

 2.5 cc. of a 2.0% heated malachite 

 green solution, prepared with sterile 

 water. 



(14) Pour 15.0 to 20.0 cc. to each Petri 

 dish. 



(15) Leave dishes open until agar is cool 

 and hard. 



Sterilization: Sterilize (10) several hours on 

 each of 2 successive days in streaming 

 steam. 



Use : Isolation of typhoid bacilli from feces. 



References : Loeffler (1906 p. 289), Klimmer 

 (1923 p. 212). 



2182. Meyer's Tissue Agar 

 Constituents: 



1. Brain infusion agar 1000.0 cc. 



2. Glucose 5.0 g. 



3. Tissue (guinea pig) 

 Preparation : 



(1) Prepare brain infusion agar (details 

 of method not given). 



(2) Dissolve 5.0 g. glucose in (1) and 

 adjust to reaction +0.5. 



(3) Boil, cool and inoculate sterile (2) 

 with edema, abdominal fluid or heart 

 blood of infected animal, properly 

 diluted. 



(4) Cut sterile (method not given) guinea 

 pig tissue into small pieces and 

 distribute them over the bottom of a 

 sterile Petri dish. 



(5) Pour (3) over tissues, and solidify agar 

 quickly on an iced glass plate. 



(6) Place plates in Novy jar, remove 

 oxygen and fill with hydrogen. 



Sterilization: Method of sterilization of 

 agar not given. 



Use: Cultivation of organism causing 

 symptomatic anthrax or black-leg in 

 swine. The author reported that other 

 tissues could not replace that of the 

 guinea pig. 



Reference: Meyer (1915 p. 467). 



2183. Williams and Burdick's Egg Veal 

 Agar 

 Constituents : 



1. Distilled water 1000.0 cc. 



2. Glycerol (15.0%) 150.0 g. ' 



3. Egg white (10.0% soln.). . . . 300.0 cc. 



4. Egg yolk (10.0%) 300.0 cc. 



5- Veal 500.0 g. 



6. NaCl 5.0g, 



7. Agar 15.0 g. 



8. Gentian violet (1.0% al- 

 coholic solution) 1.0 cc. 



Preparation : 



(1) Prepare 300.0 cc. of a 10.0% filtered 

 aqueous solution of egg white. 



(2) Prepare 300.0 cc. of a 10.0%, filtered 

 aqueous solution of egg yolk. To 

 produce the proper turbidity add 

 1.0 cc. of normal NaOH for each 

 100.0 cc. of emulsion. 



(3) Add 500.0 g. chopped lean veal to 

 1000.0 cc. of 15.0% glycerol solution 

 in water. 



(4) Infuse (3) for 24 hours. 



(5) Filter (4). 



(6) Add 5.0 g. NaCl to (5) and heat to 

 boiling. 



(7) Filter and render +1.0% alkaline. 



(8) Add 15.0 g. of powdered agar to 

 (7). 



(9) Add 1.0 cc. of a 1.0% alcoholic solu- 

 tion of gentian violet to sterile warm 

 (8). 



(10) Pour sterile (9) into sterile (1). 



(11) Add sterile (10) to sterile (2). 



(12) Mix, tube and slant. 

 Sterilization: Sterilize (1), (2) and (8) 



separately in the autoclave at 15 pounds 

 pressure for 15 minutes. 

 Use: Cultivation of B. tuberculosis. The 

 authors reported that medium contained 



