CULTURE MEDIA FOR CULTIVATION OF MICROORGANISMS 



797 



(2) Dry the ends of the eggs gently in a 

 flame. 



(3) Pierce the ends with a burned sharp 

 forcep. The end which is to be blown 

 into should be smaller and the mem- 

 brane to be left unbroken. 



(4) Blow the contents of the egg into a 

 flask containing 250.0 cc. of 5.0% 

 glycerol bouillon, neutral or mod- 

 erately alkaline to litmus. 



(5) Mi.x by a twirling motion or by gently 

 stirring with a sterile glass rod. 

 Bubbling is to be sedulously avoided. 



(6) Strain thru a cheese cloth by gravity 

 and tube. 



(7) Inspissate the tubes at 70°C. for 2 to 

 2j hours, in a moist chamber. 



(8) Incubate one week to insure sterility 

 of media. 



Sterilization: Sterilization given under 

 preparation. 



Use : Cultivation of tubercle bacilli. Park 

 and Krumwiede sealed the tube with 

 cork alone or preferably dipped the 

 cotton stopper in paraffin and then 

 stoppered with cork. Waksman studied 

 the metabolism of actinomycetes on a 

 similarly prepared medium. 



Variants : 



(a) Waksman used the following medium: 



(1) Mix 400.0 to 600.0 cc. egg and 

 1000.0 cc. bouillon containing 5.0% 

 glycerol. 



(2) Tube, slant and coagulate. 



(3) Sterilize in Arnold at 75° to 90° 

 for 3 consecutive days. 



(b) Giltner prepared the medium as 

 follows: 



(1) Wash thoroly 4 eggs with water 

 and wash in 5.0% phenol. 



(2) Allow the eggs to get partially 

 dry. 



(3) Dry the ends of the eggs in a flame. 



(4) Break the ends with a pair of 

 sterile forceps, breaking the mem- 

 brane in one end and leaving it 

 intact in the other. 



(5) Expell the contents of the eggs by 

 blowing in the end with the 

 unbroken membrane into sterile 

 Erlenmeyer flasks. 



(6) Break the yolks with a sterile wire 

 platinum and thoroly mix the 

 whites and yolks. 



(7) Add 25.0 cc. of distilled water. 



(8) Strain thru a sterile cloth. 



(9) Distribute into sterile test tubes. 



(10) Allow the tubes to stand in water 

 for 10 to 15 minutes at 45°C. to 

 drive out the air. 



(11) Slant in an inspissator and expose 

 to 70° to 75°C. for 4 or 5 hours on 2 

 successive days. 



(12) Gradually raise the temperature 

 above this on the third day and 

 finish sterilization by a single 

 exposure at 100°C. in an Arnold 

 sterilizer. 



(13) Before inoculation add 2 or 3 

 drops of sterile broth to each tube. 



(14) If human tubercle bacilli are to be 

 cultivated, add 5.0% glycerol. 



References: Park and Krumwiede (1910 p. 

 215), Waksman (1919 p. 307), Giltner 

 (1921 p. 368), Klimmer (1923 p. 223), 

 Park, Williams and Krumwiede (1924 p, 

 120). 



2447. Corper's, et al., Gentian Violet Egg 

 Medium 



Constituents: 



1. Water 1000.0 cc. 



2. Glycerol 200.0 g. 



3. NaCl 20.0 g. 



4. Beef extract (Liebig's) 12.0 g. 



5. Peptone (Witte or Difco).. . 40.0 g. 



6. Eggs 3000.0 cc. 



7. Gentian violet 1:10,000 



Preparation : 



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



(2) Adjust reaction to neutral to litmus. 



(3) Mix whites and yolks of eggs. 



(4) Mix i volume of (2) and 3 volumes of 

 (3). 



(5) Add gentian violet (Grubler's) 

 1:10,000. 



Sterilization: Inspissate or autoclave to 



sterilize, 10-15 pounds for 30 to 60 



minutes. 

 Use: Cultivation of tubercle bacilli from 



sputum. 

 Reference : Corper, Fiala and Kallen (1918 



p. 270). 



2448. Twort's Bacteria Infusion Egg 

 Medium 



Constituents : 

 1. Infusion solution. 



