CULTURE MEDIA FOR CULTIVATION OF MICROORGANISMS 



601 



(3) Dissolve 3, 4, and 5 in 900.0 cc. dis- 

 tilled water 



(4) Add (2) to (3). 



(5) Tube. 



(6) Add 2.0 to 3.0 cc. of a mixture of 

 equal parts of sterile rabbits blood 

 and citrate saline solution to 4.0 cc. 

 of the sterile agar jelly. 



(7) Heat the mixture to 45°C. for 30 

 minutes. 



(8) Slant, and allow to stand until a 

 considerable water of condensation 

 has collected, before inoculation. 



Sterilization: Sterilize (5) method not 

 given. 



Use: Cultivation of trj^panosomes. 



Authors reported that medium afforded 

 rapid growth and the vitality of the 

 cultures were not diminished. Gates 

 cultivated meningococci on a similar 

 medium. 



Variants : Gates prepared a similar medium 

 as follows: 



(1) Prepare a veal infusion agar. 



(2) Add 1.0% glucose to (1). 



(3) Adjust to pH = 7.4. 



(4) Sterilization not specified. 



(5) Just before tubing add 5.0% sterile, 

 unheated citrated horse plasma. 

 (Fresh rabbit serum may be em- 

 ployed instead of plasma.) 



References: Wolbach, Chapman and 

 Stevens (1915-16 p. 109), Gates (1919 p. 

 322). 



1899. Noguchi's Serum Plasma Agar 

 Constituents: 



1. Ringer's solution. . . 300.0 cc. 



2. Serum rabbit 100.0 cc. 



3. Plasma, citrated, 



rabbit 50.0 cc. 



4. Agar (2.0%) 5.0 to 100.0 cc. 



Preparation : 



(1) Prepare 2.0% agar. 



(2) INIix one part rabbit serum, three parts 

 Ringer's solution (see medium 180), 

 one-half part citrated rabbit plasma 

 and one-half to one part neutral or 

 slightly alkaline (1). The agar is to 

 be melted and cooled to 60 to 65°C. 

 before mixing with the other 

 materials. 



Sterilization: Not specified. 



Use: Isolation of Spirochaeta icterohaemr- 

 rhagiae. Inoculate with suspected 

 blood (material) and cover with a thin 

 layer of sterile paraffin oil. Author 

 reported that growth took place at any 

 temperature from 10°C. to 37°C. Growth 

 was nearly invisible and about 1.0 to 1.5 

 cm. below the surface. 



Reference: Noguchi (1917 p. 761). 



1900. Sparkar's Citrated Blood Agar 

 (Listen) 



Constituents: 



1. Water 2000.0 cc. 



2. Meat 1000.0 cc. 



3. Peptone 40.0 g. 



4. NaCl 20.0 g. 



5. Agar 60.0 g. 



6. Blood (human) 

 Preparation : 



(1) Boil 1000.0 g. of meat in 2 liters of 

 water for 40 minutes. 



(2) Filter. 



(3) Dissolve 3, 4, and 5 in sterile (2). 



(4) Draw one volume of human blood 

 into a flask containing 4 volumes 

 sterile citrated saline (0.5% sodium 

 citrate and 0.85% NaCl). 



(5) Mix the blood and saline thoroly. 



(6) Heat in a water bath at 65°C. for 30 

 minutes. 



(7) Remove precipitate by filtering thru 

 sterilized filter paper. 



(8) Add 1.0 cc. of (7) to 10.0 cc. of melted 

 (3) at 50°C. 



(9) Slant. 



Sterilization: Sterilize (2) at 15 pounds 



pressure in the autoclave 

 Use: Cultivation oi B . influenzae (Pfeiffer). 

 Variants : 



(a) Malone prepared a similar medium as 

 follows: 



(1) Method of preparation or exact 

 composition of nutrient agar not 

 given. 



(2) Add 10.0 cc. of pigeons blood to 

 90.0 cc. of 1.0% sodium citrate 

 solution in normal salt solution. 



(3) Heat (2) for ^ hour at 66° to 70°C. 



(4) Filter thru filter paper. 



(5) Add 1.0 cc. of (4) to 10.0 cc. of 

 nutrient agar at a temperature of 

 50° to 60 °C. 



