CULTURE MEDIA FOR CULTIVATION OF MICROORGANISMS 



595 



(2) Steam for 60 minutes. 



(3) Desiccate in vacuo over sulphuric 

 acid or calcium chloride. 



(4) Grind the residue to powder. 



(5) Make for use a 10.0% solution of 

 blood powder. 



(6) When ready for use add 3 parts 

 (5) at 45°C. to 7 parts infusion 

 agar at 45 °C. 



(b) (1) Mix equal parts 12.0% crystalline 

 Na2C03 solution and defibrinated 

 blood. 

 (2) Mix 3 parts (1) with 7 parts of 

 4.0% infusion agar. 

 Reference: Harvey (1921-22 pp. 73, 75). 



1891. Filde's Blood Digest Agar 

 (Kristensen) 



Constituents: 



1. Water 25,000.0 cc. 



2. Beef 25,000.0 g. 



3. Peptone 1.0% 



4. NaCl 0.5% 



5. Blood, defibrinated sheep 

 or horse 



Preparation : 



(1) Add 6.0 cc. of pure concentrated 

 HCl to 150.0 cc. of a 0.9% NaCl 

 solution, and then 50.0 cc. defibri- 

 nated horse or sheep blood and 2.0 g. 

 Langebek's concentrated pepsin. 



(2) Pour into a sterile flask and incubate 

 at 56°C. for 5 or 6 hours. 



(3) Add NaOH solution until a sample 

 diluted with distilled water to a 

 light brown color turns red on the 

 addition of phenol red, without 

 giving the alkaline color change 

 with a-naphtholphthalein. 



(4) Add 0.25% chloroform and stopper 

 the flask with a sterile rubber and 

 keep in a cold room. 



(5) Mince 25,000.0 g. beef. 



(6) Add 25 liters of water and allow to 

 stand in the cold until the next day. 



(7) Boil for a short time. 



(8) Press the fluid from the meat. 



(9) Add about 16 liters of water to the 

 meat, heat to boiling once more and 

 again press the juice from the meat. 



(10) Mix the fluid from (7) and (8). 



(11) Add 1.0% peptone and 0.5% NaCl. 



(12) Adjust to pH about 7.8 to 7.9. 



(13) Add 2.0% agar and dissolve. 



(14) Filter thru cotton wool and distrib- 

 ute in 600.0 cc. flasks. 



(15) The pH of sterile agar should be 

 about 7.2. 



(16) Add 30.0 cc. of (4) to each 600.0 cc. 

 of melted (15) cooled to 40 to 50°C. 



Sterilization: Sterilize (10) by heating in 

 the steamer on each of 3 consecutive days. 



Use: Cultivation of influenza bacilli, 

 Haemoglobinophilic bacteria. 



Reference: Kristensen (1922 p. 229). 



1892. MacNeal's Blood Infusion Agar 



Constituents: 



1. Water 1000.0 cc. 



2. Chopped beef 125.0 g. 



3. Agar 20.0 g. 



4. Peptone 20.0 g. 



5. NaCl 5.0 g. 



6. N/1 NaoCOa soln 10.0 cc. 



7. Blood, naturally sterile de- 

 fibrinated rabbit 2000.0 cc. 



Preparation : 



(1) Extract 125.0 g. beef in 1000.0 cc. 

 distilled water. 



(2) Dissolve 3, 4, 5 and 6 in (1). 



(3) Tube. 



(4) Add to sterile (3), cooled to about 

 60° two volumes of naturally sterile 

 defibrinated rabbit's blood. Mix 

 thoroly. 



(5) Slant and solidify. 

 Sterilization: Sterilize (3) in the autoclave. 

 Use: Isolation of Nagana parasite, Tr. 



Brucei and other trypanosomes. In- 

 oculate Nagana blood in the small amount 

 of liquid which collects on the surface 

 of the blood agar. Thompson cultivated 

 trypanosomes found in gold fish (T. 

 danilewskyi) . 

 Variants : 



(a) The author used 4 times as much meat 

 (500.0 g. per liter) for succeeding 

 cultures. 



(b) Tobey used the same medium and 

 added two volumes of defibrinated 

 rabbit's blood. He inoculated the 

 water of condensation with a drop of 

 blood taken from the heart of a bird 

 by means of a drawn-out tube pipette. 

 The cotton plug is then cut short, 

 moistened with mercuric chloride 



