520 



CULTURE MEDIA FOR CULTIVATION OF MICROORGANISMS 



sterile lactic, tartaric or hydrochloric 

 acid to sterile tubes of agar. 



(b) Duval cultivated B. leprae in media 

 prepared as follows: 



(1) Method of preparation or composi- 

 position of 2.0% nutrient agar not 

 given. 



(2) Adjust to 1.5% alkaline to phenol- 

 phthalein. 



(3) Distribute in 10.0 cc. lots. 



(4) Method of sterilization not given. 



(5) Prepare 2 0% solutions of cystine, 

 (made from protein) leucine and 

 tryptophane. 



(6) Sterilize (5) by passing thru a 

 Berkefeld filter. 



(7) Add 5.0 cc. of one of (6) or in com- 

 bination to melted (4). 



(8) Mix thoroughly and solidify in a 

 slanted position. 



(c) Abel mixed one part serum, ascitic 

 fluid or hydrocele fluid, warmed, to 

 40 to 50°C. with one or two parts agar. 



(d) Tanner added 1.0% of any desired 

 carbohydrate, alcohol, etc. to plain 

 agar. 



(e) Dopter and Sacquepee added 2.0% 

 of any desired carbohydrate, alco- 

 hol, etc., to nutrient agar. 



References: Kohler (1893 p. 76), Thoinot 

 and Masselin (1902 p. 35), Duval (1910 

 p. 655), Abel (1912 p. 26), Tanner (1919 

 p. 48), Dopter and Sacquepee (1921 

 p. 128). 



1710. Wurtz's Nutrient Agar 



Constituents : 



1. Bouillon 1000.0 cc. 



2. Agar 12.0 g. 



Preparation : 



(1) Dissolve 12.0 g. of agar in a liter of 

 bouillon, slightly alkaline, by heating 

 in a salt water bath. 



(2) Cool to 50°C. and add the whites of 

 two eggs beaten up in a liter of 

 water. 



(3) Heat for one minute at 125°C. 



(4) Filter thru Chardin filter paper. 



(5) Distribute in tubes. 

 Sterilization: Sterilize at 115°C. for 



15 minutes. 

 Use : General culture medium. 

 Variants: The authors listed below have 



given the following method of prepara- 

 tion for similar media: 

 (a) Smith (1902). 



(1) Grind 5.0 g. of agar and boil for one 

 or two hours in 100.0 cc. of water in 

 a beaker. Add water from time 

 to time. 



(2) Add 250.0 cc. of meat infusion 

 (or extract) peptone solution. 



(3) Boil until the weight of the bouil- 

 lon plus 5.0 g. of agar is obtained. 



(4) Cool to 60°C. or less and add one 

 half the white of an egg dissolved 

 in 25.0 cc. of boiled water. 



(5) Reboil for 10 or 15 minutes to 

 coagulate the egg albumin. 



(6) Filter thru a folded and moistened 

 paper. 



(7) Make up to the weight of the 

 bouillon plus 5.0 g. agar. 



(8) If the agar is not clear, reheat 

 and refilter. 



(9) Distribute. 



(10) Sterilize intermittently or in the 

 autoclave. 



(b) Roux and Rochaix (1911). 



(1) Dissolve 15.0 g. of agar in a liter of 

 peptone bouillon by heating at 

 100°C. 



(2) Filter thru muslin. 



(3) Cool to 70-75°C. and add the white 

 of an egg. Mix well. 



(4) Readjust the reaction if necessary. 



(5) Heat for 45 minutes. 



(6) Filter while hot. 



(7) Distribute in tubes. 



(8) Sterilize in the autoclave, 



(c) Roux and Rochaix (1911). 



(1) Soak 25.0 g. of chopped agar for 

 24 hours in 500.0 cc. of water, 

 acidulated by the addition of 6.0% 

 HCI. Stir occasionally. 



(2) Wash thoroly with water. 



(3) Soak the agar for 24 hours in 

 500.0 cc. of a 5.0% ammonia 

 solution. 



(4) Wash thoroly. 



(5) Place in a liter of bouillon and 

 heat until dissolved. 



(6) Neutralize by the addition of a 

 saturated solution of NaHCOs. 



(7) Pass thru flannel and then filter 

 using a hot water funnel. 



(8) Distribute into flasks or tubes. 



