324 



CULTURE MEDIA FOR CULTIVATION OF MICROORGANISMS 



Tryptophane Broth in hot distilled 

 water. 

 (2) If sterilized at 15 pounds pressure for 

 20 minutes pH = 6.8±. 

 Sterilization: Sterilize at 15 pounds pres- 

 sure for 20 minutes. 

 Use: Indol production. Medium permits 



a strong positive indol test in 12 hours. 

 Reference: Digestive Ferments Co. (1925 

 p. 13). 



1128. Bacto Peptonized Milk (Dehydrated) 



Constituents : 



1. Distilled water 1000.0 cc. 



2. Bacto Peptonized Milk 

 (Dehydrated) 15.0 g. 



Preparation: 



(1) Dissolve 15.0 g. Bacto Peptonized 

 Milk (Dehydrated) in 1000.0 cc. dis- 

 tilled water. 



(2) If sterilized for 20 minutes at 15 

 pounds pressure pH = 6.5zt. 



Sterilization: Sterilize at 15 pounds pres- 

 sure for 15 minutes. 



Use: Cultivation of lacto-bacillus group. 



Variants: Add 0.1% Bacto agar to medium 

 for maintaining stock cultures. 



Reference: Digestive Ferments Co. (1925 

 p. 14). 



1129. Mueller's Trypsinized Casein Solution 



Constituents: 



1. Water 1000.0 cc. 



2. NaCl (1.0%) 10.0 g. 



3. MgS04 (0.04%) 0.4 g. 



4. CaCl, (0.02%) 0.2 g. 



5. K2HPO4(0.2%) 2.0 g. 



6. Glucose (0.2%) 2.0 g. 



7. Phenol red (0.02% solution). 80.0 cc. 



8. Casein 

 Preparation : 



(1) Dissolve 2, 3, 4, 5, 6 and 7 in 1. 



(2) Digest a solution of commercial casein 

 with Fairchild's trypsin at 37°C. for 

 2 weeks. 



(3) Boil. 



(4) Filter. Twelve cubic centimeters of 

 the filtrate represents about 0.5 g. of 

 casein. This is an impure casein 

 digest. 



(5) Mix 12.0 cc. of (4) with 13.0 cc. of 

 water and add 25.0 cc. of (1). 



(6) Adjust to pH = 8.0. 



Sterilization: Sterilize in autoclave at 10 

 pounds for 10 minutes. 



Use: To study requirements of strepto- 

 cocci and pneumococci. Author reported 

 that the organisms grew in impure casein 

 medium and not in pure casein. 



Variants: The author prepared a pure 

 casein digest by precipitating a solution 

 of commercial casein in Na2C03 three 

 times by acetic acid. Then washed pre- 

 cipitate in alcohol and ether. Treat this 

 purified casein as in steps (2), (3), (4) and 

 (5) above. 



Reference: Mueller (1922 pp. 318, 319). 



1130. Cole and Onslow's Trypsinized Casein 

 Solution 



Constituents: 



1. Distilled water 1000.0 cc. 



2. Casein 200.0 g. 



3. NaCl (0.5% soln.) 2000.0 cc. 



Preparation : 



(1) Obtain the fresh pancreas of a pig, 

 free it from fat as far as possible 

 and weigh. 



(2) Mince (1) finely and add 3 times its 

 weight of distilled water, and its own 

 weight of strong alcohol. 



(3) Shake the mixture well in a large 

 bottle, and allow to stand for 3 days 

 at room temperature, shaking occa- 

 sionally. 



(4) Strain thru muslin and filter thru a 

 large folded filter paper. 



(5) Measure the filtrate and add 1.0 cc. 

 of concentrated HCl for each liter 

 of the filtrate. 



(6) Allow to stand for a few days and 

 filter off the sediment. 



(7) The filtrate may be stored in a stop- 

 pered bottle, without the addition 

 of an antiseptic. If the extract is to 

 be used at once it is not necessary 

 to add the HCl. 



(8) Add 20.0 g. of anhydrous NazCOa to 

 one liter of tap water, and boil in a 2 

 liter flask or large evaporating basin. 



(9) When (8) is boiling, transfer the 

 vessel to a boiling water bath, and 

 gradually dust in 200.0 g. of casein 

 (Laitproto j^6 for bacteriological 

 purposes from Casein Limited). 

 Stir well to avoid the formation of 

 lumps. 



