CULTURE MEDIA FOR CULTIVATION OF MICROORGANISMS 



665 



(9) Filter thru muslin to remove fat. 



(10) Add 1.0 cc. glacial acetic acid per 

 pot. 



(11) Sterilize by heating up to 110°C. 

 and then cool. 



(12) Filter to remove mince. 



(13) Make slightly alkaline to litmus and 

 then add 2.5 g. NaCl and 0.125 g. 

 CaCl2 per liter. 



(14) Sterilize at 115°C. for one hour. 



(15) Filter thru paper. 



(16) Put 100.0 cc. broth into each whiskey 

 bottle. 



(17) Sterilize at 120°C. for one hour. 



(18) Add 4.0 g. desiccated agar to each 

 bottle. 



Sterilization: Sterilize (18) at 120°C. for 

 one hour. 



Use: Growth of organisms for producing 

 vaccines. The quantities employed 

 yielded about 15 or 16 bottles. Authors 

 reported this medium inferior to medium 

 2052 for vaccine production. 



Reference: Norris (1919-20 p. 542). 



2063. Harvey and Iyengar's Desiccated 

 Tryptonized Mutton Bouillon Agar 



Constituents: 



1. Trypsinized mutton 1000.0 cc. 



2. Agar 

 Preparation : 



(1) Prepare a tryptic digest of mutton 

 bouillon (method not given). 



(2) Adjust (1) to pH = 8.0. 



(3) Cut up agar fiber into small pieces. 



(4) Add (3) to (2). (6.0% by weight.) 



(5) Autoclave for one hour at 120° to 

 melt the agar. 



(6) Filter thru cotton, wool and muslin 

 into a tin receptacle. 



(7) Cut the agar out of the receptacle 

 and into slices. 



(8) Paes slices thru meat mincing ma- 

 chine with a finely perforated outlet 

 disc. 



(9) Spread minced nutrient agar on metal 



or other type of trays. 



(10) Dry in hot air oven or any other con- 

 venient way. 



(11) Store powder in sterile glass stop- 

 pered bottle. 



(12) Add 4.0% to 6.0% by weight of (11) 

 to water in test tubes or flasks. 



Sterilization: Keep (11) immersed for 1 to 

 2 hours in boiling water or in a steam 

 sterilizer to bring the medium into 

 solution and to sterilize. 



Use: Desiccated nutrient medium. 



Variants: Vardon prepared a similar me- 

 dium as follows: 



(1) Wash a quantity of agar fiber in tap 

 water containing 0.25% acetic acid. 



(2) Wash in several changes of tap water 

 until the washings are neutral to lit- 

 mus paper. 



(3) Place the washed fiber on perforated 

 or wire gauze trays and cover with 

 thin muslin. 



(4) Expose the agar in the trays to the 

 sun to dry. 



(5) Prepare a double strength^ (composi- 

 tion not given) tryptic digest of 

 mutton. 



(6) Filter thru paper and measure the 

 filtrate. 



(7) Adjust the reaction of the filtrate to 

 pH = 8.0. 



(8) Add2 after this adjustment 25.0% 

 more alkali of the strength used. 



(9) Add 9.0%3 by weight of washed and 

 dried agar fiber. 



(10) Place the mixture for one hour in 

 the autoclave, at 120 °C. to thoroly 

 dissolve the agar. 



(11) Allow the nutrient agar to solidify^ 

 in a metal funnel. 



(12) Turn out the clear solidified mass 

 and cut into pieces. 



(13) Pass thru a meat mincing machine 

 with finely perforated outer disc. 



(14) Spread the minced agar on metal 

 trays. 



(15) Dry in hot air oven at a tempera- 

 ture not exceeding* 60°C. 



(16) Remove as soon as the agar is thoroly 

 dried. 



(17) Scrape the dried material off the 

 trays and grind to powder. 



(18) Replace the powder in the hot oven 

 and complete the drying process. 



(19) Record the final^ weight of the 

 powder. 



(20) Store in tight stoppered bottles. 

 Notes: ^All the constituents of 

 the ordinary tryptic digest bouillon 

 (Douglas' medium) are doubled, 

 i.e. the meat, the liquid pancreatic 



