CULTURE MEDIA FOR CULTIVATION OF MICROORGANISMS 



239 



Variants : Harvey added one of the follow- 

 ing combinations of dyes or indicators to 

 a liter of meat infusion peptone solution 

 prepared as indicated in variant (bb) 634. 



(a) Cresol red (1.6% solution in 95.0% 

 alcohol) 1.0 cc. 



(b) Brom cresol purple (1.6% solution in 

 95.0% alcohol) 4.0 cc. 



(c) Brom cresol purple, cresol red indi- 

 cator solution 1.0 cc. 



(d) China blue 0.025 g. + rosolic acid or 

 its sodium salt 0.05 g. 



(e) China green (0.2% solution) 15.0 cc. 



(f) Malachite green 1.0 g. 



(g) China blue 0.025 g. + phenol red 

 (phenol sulphone phthalein) 0.01 g. 



Reference: Muller (1899 p. 805), Harvey 

 (1921-22 pp. 88-92), Omelianski (1903 

 p. 4). 



785. Robinson and Rettger's Infusion 

 Opsine Solution 



Constituents: 



1. Beef Infusion 1000.0 cc. 



2. Opsine 20.0 g. 



Preparation : 



(1) Method of preparation or composi- 

 tion of beef infusion not given. 



(2) Dissolve 2 in (1). 



(3) Adjust reaction faintly alkaline to 

 litmus. 



(4) Distribute in 20.0 cc. lots in 150.0 cc. 

 Erlenmeyer flasks. 



Sterilization: Sterilize at 12 to 14 pounds 

 pressure for 15 minutes. 



Use: Production of diphtheria toxin by 

 B. diphtheriae. The author reported 

 that the addition of 0.1% glucose had 

 little effect. M.L.D. = 0.035 cc. 



Variants : 



(a) The author added 0.1% glucose with 

 or without 0.5 g. XaCl. 



(b) The author added 0.5 g. XaCl. 

 Reference: Robinson and Rettger (1917 



p. 367). 



786. Warden, Connell and Holly's Veal 

 Infusion Broth 



Constituents : 



1. Veal Infusion 1000.0 cc. 



2. Peptone (Difco proteose)... 20.0 g. 



3. NaCl 5.0 g. 



Preparation : 



(1) Method of preparation of veal infu- 

 sion not given. 



(2) Dissolve 2 and 3 in 1. 



(3) Adjust to pH = 7.9 by the addition 

 of XaOH. 



Sterilization: Sterilize in the autoclave. 



Use: Cultivation of C. diphtheriae and 

 B. megatherium and to produce large 

 quantities of toxin. Authors reported 

 that a very high grade of toxin was 

 produced. 



Variants : The following authors have sug- 

 gested various methods of preparing this 

 medium: 



(a) Dernby and David. 



(1) Immerse 6.0 g. of fresh finely 

 chopped veal in 12 liters of tap 

 water and add 70.0 g. of ordinary 

 distillers yeast. 



(2) Incubate at 36°C. for 16 hours. 



(3) Press thru cheese cloth. 



(4) Add 1.5% Witte's peptone and 

 0.4% NaCl. 



(5) Adjust to pH = 7.2 or 7.3. 



(6) Boil several minutes. 



(7) Filter while warm thru paper. 



(8) Distribute in liter flasks. 



(9) Sterilize at 110° for 20 minutes. 



(b) Dernby and Blanc. 



(1) Chop veal finely and allow to 

 autolyzeat37°for24hours. Then 

 boil and filter. 



(2) Dissolve 2.5 g. XaCl and 5.0 g. 

 peptone in (1). 



(3) Adjust to desired pH with XaOH 

 and HCl. 



(4) Sterilize at 110° for 20 minutes. 



(c) Harvey. 



(1) Mince finely fat-free veal. 



(2) Add 500.0 g. to 1000.0 cc. tap water. 



(3) Add an 18 hour culture of B. coli. 



(4) Incubate 20 hours at 22°C. 



(5) Heat the mixture 2 hours at a 

 temperature not exceeding 50°C. 



(6) Skim off fat on the surface. 



(7) Raise to boiling point. 



(8) Boil 10 minutes. 



(9) Pour the mixture onto a wet, thick, 

 clean cloth. 



(10) Collect the fluid which drains thru 

 the cloth together with that ob- 

 tained by squeezing the meat in 

 the cloth. 



(11) Filter the fluid collected thru well 

 wetted, thick filter paper. 



(12) Add to the filtrate: peptone 

 20.0 g., sodium chloride 5.0 g. 



