794 



CULTURE MEDIA FOR CULTIVATION OF MICROORGANISMS 



(5) Sterilize 20 minutes at 100°C. 3 

 days, 24 hours after completion of 

 coagulation. 



(6) Test sterility before use. 



(d) Harvey also added 1.0 cc. glycerol to 

 each test tube before final steriliza- 

 tion to convert into a medium suit- 

 able for cultivation of B. tuberculosis 

 The addition of a few drops of 1.0% 

 neutral red gives a color to the 

 medium which serves to show up 

 colonies, and serves as an indicator 

 of the production of acid. 



(e) Stitt cultivated tubercle bacilli on 

 the following medium: 



(1) Prepare a 1.0% glucose bouillon. 



(2) Flame a mortar wet with alcohol. 



(3) Crack a clean egg with a sterile 

 knife and allow the contents to 

 drop into the sterile mortar. 



(4) Add from 10.0 to 15.0 cc. of sterile 

 (1) to (3). 



(5) Mix into a smooth paste. 



(6) Inspissate for 2 hours in the rice 

 cooker. If the mortar and knife 

 are not sterilized it is necessary to 

 sterilize after inspissation. 



(7) One cc. of glycerol bouillon may be 

 added to each tube before final 

 sterilization. 



(8) Plug the tubes with a rubber cap, 

 or preferably, heat the end of the 

 tube, withdraw the plug quickly 

 and dip the part of the plug that 

 enters the tube into hot melted 

 paraffin. Then quickly reintroduce 

 the plug. 



References: Roddy (1917 p. 43), Harvey 

 (1921-22 p. 85), Stitt (1923 p. 42). 



2442. Putnam and Gay's Milk Egg Medium 



Constituents : 



1. Litmus milk 200.0 cc. 



2. Egg 100.0 cc. 



3. Glycerol (5.0%) 15.0 cc. 



4. Peptone (0.8%) 2.4 g. 



5. Beef extract (0.4%) 1.2 g. 



6. Glucose (1.2%) 3.6 g. 



Preparation : 



(1) Mix one part egg with two parts lit- 

 mus milk. 



(2) Add 5.0% glycerol, 0.8% peptone, 

 0.4% beef extract and 1.2% glucose. 



(3) Adjust to 1.14% acid— indicator not 

 specified. 



(4) Inspissate and sterilize in the Arnold. 

 Sterilization: See step (4) above. 



Use: To determine growth of influenza 

 bacilli in mi.xed cultures. Author re- 

 ported that medium does not support the 

 growth of the influenza bacillus in mixed 

 culture. 



Reference: Putnam and Gay (1920-21 p 4). 



2443. Petroff's Gentian Violet Egg Medium 



Constituents : 



1. 15.0% glycerol in water. 500.0 cc. 



2. Beef (or veal) 500.0 g. 



3. Whole egg 1000.0 cc. 



4. Gentian violet 1 to 10,000. 



Preparation : 



(1) Infuse 500.0 g. beef or veal in 500.0 

 cc. of a 15.0% solution of glycerol in 

 water for 24 hours. 



(2) Squeeze in a sterile meat press and 

 collect in a sterile beaker. 



(3) Sterilize the shells of eggs by im- 

 mersing for 10 minutes in 70.0% 

 alcohol or by pouring hot water 

 upon them. 



(4) Break the eggs into a sterile beaker 

 and mix. 



(5) Filter thru a sterile gauze. 



(6) Add two parts by volume of (5) to 

 one of (2). 



(7) Add sufficient 1.0% alcoholic gentian 

 violet to make a dilution of 1 to 

 10,000. 



(8) Tube in 3.0 cc. lots in sterile test 

 tubes and inspissate for 3 successive 

 days, 85°C. the first day. On the 

 second and third days for not more 

 than one hour at 75°C. 



Sterilization: Method not given. 

 Use: Isolation and cultivation of tubercle 

 bacilli. For the cultivation of bovine 

 tubercle bacilli, the meat was infused in 

 plain water, glycerol not being added. 

 Waksman cultivated actinomyces on a 

 medium similarly prepared. 

 Variants: The following investigators pre- 

 pared the medium as follows: 

 (a) Keitly prepared the medium like 

 above. To sterilize, however, he 

 placed the tubes in the inspissator. 

 Bring the temperature to 95°C. as 

 quickly as possible, avoiding bubbles. 



