230 



CULTURE MEDIA FOR CULTIVATION OF MICROORGANISMS 



(i) Smith (1902). 



(1) Soak 500.0 g. of finely chopped 

 fresh fat free beef in 1000.0 cc. 

 of boiled water over night in the 

 ice box. 



(2) Strain thru a piece of cheese 

 cloth. Compress the pulp to 

 obtain a liter of fluid. Make up 

 to a liter if necessary. 



(3) Heat between 50° and 60°C. and 

 add 10.0 g. peptone and 5.0 g. 

 NaCl. Stir constantly until solu- 

 tion is complete. 



(4) Boil with constant stirring. 



(5) Filter thru paper. 



(6) Reboil and filter again if the 

 filtrate is turbid. 



(7) Cool to room temperature and 

 make the volume to one liter. 



(8) Adjust the reaction. 



(9) Boil and filter. 



(10) Tube or flask. 



(11 ) Sterilize by the fractional method 

 in steam at 100°C. or in the auto- 

 clave at 120°C. 



(j) Frost (1903). 



(1) Remove all the fat and connec- 

 tive tissue from 500.0 g. of beef 

 and mince (or use hamburg 

 steak). 



(2) Add 1 liter of distilled water to 

 (1), shake thoroughly and set in 

 the ice box for 12 to 24 hours. 



(3) Squeeze thru a cloth and add 

 enough distilled water to make 

 1 liter. 



(4) Add 1.0% peptone (Witte) and 

 0.5% NaCl to (3). 



(5) Weigh the solution and vessel. 



(6) Heat to not above 60°C. until 

 the ingredients are in solution. 



(7) Restore the loss in weight by the 

 addition of distilled water. 



(8) Neutralize to phenolphthalein 

 by the addition of normal NaOH. 



(9) Boil for 5 minutes and restore the 

 weight. 



(10) Test the reaction and readjust if 

 necessary. 



(11) Add 0.5 to 1.5% of normal HCl. 



(12) Heat until the precipitate ap- 

 pears flaky. 



(13) Filter thru moistened filter paper. 



(14) Tube. 



(15) Sterilize for 20 minutes in a 

 steam sterilizer on 3 consecutive 

 days or in the autoclave at 120°^ 

 for 20 minutes. 



(k) Frost (1903). 



(1) Remove all the fat and con- 

 nective tissue from 500.0 g. of 

 beef and mince or use hamburg 

 steak. 



(2) Add 1 liter of distilled water. 



(3) Place in a vessel for cooking and 

 cook for 30 minutes at about 

 70°C. 



(4) Filter thru paper and make up 

 to one liter. 



(5) Add 1.0% peptone (Witte) and 

 0.5% NaCl to (3). 



The remainder of the preparation 

 is identical with variant (j) 

 above, from step (5). These two 

 bouillons may be inoculated with 

 B. coll and incubated at 38 °C. 

 for several hours to render them 

 sugar free. 



(I) Committee A. P. H. A. (1905). 



(1) Infuse 500.0 g. finely chopped 

 lean meat with 1000.0 cc. water 

 for 24 hours in refrigerator. 



(2) Make up volume. 



(3) Strain thru cotton flannel. 



(4) Add 10.0 g. Witte's peptone. 



(5) Dissolve by heating in water 

 bath. 



(6) Heat over steam bath 30 minutes. 



(7) Restore volume. 



(8) Adjust reaction to +1 with 

 phenolphthalein. 



(9) Boil 2 minutes over free flame, 

 stir. 



(10) Restore volume. 



(II) Filter thru absorbent cotton and 

 cotton flannel until clear. 



(12) Tube in 10.0 cc. lots. 



(13) Sterilize— autoclave or by inter- 

 mittent method. 



(m) Heinemann (1905). 



(1) Add 500.0 g. lean minced beef, 

 free from fat and tissue as pos- 

 sible, to 1000.0 cc. tap water. 



(2) Place on ice for 24 hours. 



(3) To prepare a sugar free broth 

 add a pure culture of Bacillus coli 

 to (2) and incubate at 37° for 

 24 hours. 



