672 



CULTURE MEDIA FOR CULTIVATION OF MICROORGANISMS 



(1) Free horse meat from all fat and 

 tendons. 



(2) Chop into small pieces with a 

 knife, a scraper or meat grinding 

 machine until there are only very 

 loose particles. 



(3) To 125.0 g. of (2) add 3.0 g. of 

 fresh Witte-Rostock pepsin 400.0 

 cc. distilled water and 2.0 cc. of a 

 50.0% HCl solution. 



(4) Place in an Erlenmeyer flask and 

 incubate at 37 °C. 



(5) Incubate for 2 days, shaking oc- 

 casionally and adjusting the re- 

 action with the addition of a 50.0% 

 HCl solution. 



(6) Filter the grey white precipitate 

 which has formed at the end of 

 this time. 



(7) Test a portion of the filtrate by 

 the Biuret reaction (violet colora- 

 tion with KOH and CUSO4) which 

 must be positive. 



(8) Add 3.9 g. of NaaCOs (siccum) to 

 the filtrate. 



(9) Sterilize (Method not given). 



(10) Cut a hog's pancreas into small 

 pieces with a knife, and place in 

 the ice box for 24 hours. 



(11) Add 40.0 cc, of glycerol and 160.0 

 cc. distilled water, and extract for 

 several days in the ice box. 



(12) Press out the juice and add a small 

 piece of camphor to the liquid. 



(13) Add 15.0 cc. of (12) to (9) by means 

 of a sterile pipette. 



(14) Incubate at 37°C. for six hours. 



(15) Sterilize immediately in the auto- 

 clave, and neutralize with 50.0% 

 HCl. 



(16) Add 1950.0 cc. of water, 6.0 g. 

 NaCl, 39.0 agar to (15). 



(17) Boil in the autoclave for 3 hours. 



(18) Filter thru cotton and distribute 

 into little flasks. 



(19) Sterilize as usual (exact method 

 not given). 



(b) Bosse (Kolle and Wasserman) pre- 

 pared the medium as follows: 

 (1) Mix 150.0 g., of finely chopped 

 horse heart with 3.0 g. of Witte's 

 pepsin (fresh) 2.0 cc. of 50.0% 

 HCl and 400.0 cc. of distilled 

 water. 



(2) Digest for two days at 37°C. 



(3) Filter, and if positive to the 

 Biuret test add 3.9 g. dry NajCOs. 



(4) Add 15.0 cc. of a glycerol extract 

 of a hog's pancreas to (3). 



(5) Incubate for 6 hours at 37°C. 



(6) Sterilize in the steamer. 



(7) Neutralize with HCl. 



(8) Add 1950.0 cc. water, 6.0 g. NaCl 

 and 39.0 g. agar to (7) and boil for 

 3 hours in the steamer. 



(9) Filter thru cotton. 



(10) Distribute in flasks. 



(11) Sterilize. 



(c) Harvey gave the following method of 

 preparation: 



(1) Mix 150.0 g. of finely minced horse 

 heart, 5.0 g. pepsin, 2.0 cc. of 

 50.0% HCl and 400.0 cc. of distilled 

 water. 



(2) Leave to digest 2 days at 37 °C 



(3) Filter. 



(4) Add 40.0 cc. of glycerol and 160.0 

 cc. distilled water to a finely 

 chopped pig pancreas. 



(5) Infuse (4) for 3 days in an ice chest, 

 adding a small piece of camphor. 



(6) Add 3.9 g. anhydrous NaaCOj and 

 15.0 cc. of (5) to the filtrate from 

 (3). 



(7) Leave 6 hours at 37°C. 



(8) Sterilize at 100°C. 



(9) Neutralize with HCl. 



(10) Add 1950.0 cc. water, 6.0 g. sodium 

 chloride, and 39.0 g. agar. 



(11) Steam 3 hours. 



(12) Filter. 



(13) Distribute into test tubes and 

 flasks. 



(14) Sterilize. 



(d) Deycke (Klimmer) gave the follow- 

 ing method of preparation : 



(1) Mix 125.0 g. of finely chopped 

 horse meat with 400.0 cc. distilled 

 water and 3.0 g. of Witte's pepsin. 



(2) Add 2.0 cc. of 50.0% concentrated 

 HCl. 



(3) Incubate at 37°C. for 48 hours, 

 adding more HCl if necessary. 



(4) Filter. 



(5) Add 3.9 g. water free soda to the 

 filtrate. 



(6) Sterilize (method not given). 



(7) Chop the pancreas of a hog into 



