CULTURE MEDIA FOR CULTIVATION OF MICROORGANISMS 



671 



lowish green and contains very little 

 undigested debris. 



(4) Transfer to large bottles and steam 

 for 10 minutes to stop digestion. 



(5) Cool to 80°C. and make faintly alka- 

 line to litmus using 2N KOH or 2N 

 NajCOs. 



(6) Cool to 37°C. and add 1.0% pan- 

 creatic e.xtract or "Bacto" trypsin. 



(7) Keep the mixture at 37°C. for 3 to 

 10 hours depending on the action of 

 the trypsin and the digestion de- 

 sired. Control the process by re- 

 peated tests for tryptophane. 



(8) When trypsinizing is sufficiently ad- 

 vanced render slightly acid with 

 glacial acetic acid, and bring slowly 

 to boiling point for 10 minutes. 



(9) Filter thru paper or keep in cool 

 place over night and decant the 

 clear liquid in the morning. 



(10) Add agar and K2HPO4 and autoclave 

 at 10 pounds pressure for 45 minutes 

 or heat in double boiler to lOO^C. 

 until the agar is dissolved. 



(11) Restore the volume lost by eva- 

 poration. 



(12) Adjust to slightly alkaline to litmus 

 or pH = 7.3 by using 2N KOH or 

 NaOH. 



(13) Cool to 60°C. and add white of egg 

 with crushed shells or for sake of 

 economy, ordinary beef or sheep 

 serum, in the quantity of 25-50.0 cc. 

 per liter. 



(14) Autoclave for one hour at 115''C. 



(15) Filter thru cotton and distribute in 

 200-500.0 cc. quantities. 



Sterilization: Sterilize at 100° for 30 min- 

 utes on two successive days or at 

 10 pounds pressure for 15 minutes. 



Use: General inexpensive culture medium. 



Variants: The authors prepared a sugar- 

 free medium as follows: 

 (1) Carry out procedure as above steps 

 (1) thru (5). 



(6) Cool to 37°C. and add 1.0% pan- 

 creatic extract or Bacto trypsin, 

 8.0 g. K2HPO4 and 1.0% of a culture 

 of B. saccharolyte. 



(7) Keep the mixture at 37°C. for 3 to 

 10 hours depending on the action of 

 the trypsin and digestion desired. 

 Control the process by repeated tests 

 for tryptophane. 



(8) When the digest is sugar free add 

 4.0 g. CaCOs and 80.0 g. agar. 



(9) Autoclave at 10 pounds pressure for 

 45 minutes or heat in double boiler to 

 100°C. until the agar is dissolved. 

 Remainder of the preparation same 

 as medium given above. 



Reference: Stickel and Meyer (1918 p. 80). 



2074. Deycke and Voigtlander's Tryptic and 

 Peptic Meat Digest Agar 



Constituents : 



1. Distilled water to 1950.0 cc. 



2. Meat, horse 125.0 g. 



3. HCl (5.0%) 2.0 cc. 



4. NaoCOa (siccum) 3.7 g. 



5. Pancreatin (Merck's gly- 

 cerol) 15.0 cc. 



6. NaCl 6.0 g. 



7. Glycerol 



8. Agar 

 Preparation: 



(1) Grind up 125.0 g. of lean horse meat 

 with 3.0 g. pepsin and 400.0 cc. 

 distilled water with 2.0 cc. of HCl. 



(2) Place in an Erlenmeyer flask and 

 incubate at 37°C. until there is 

 complete solution. 



(3) Neutralize with NaaCO 3 (anhydrous) 

 and divide into 3 portions. 



(4) Sterilize each portion (exact method 

 not specified). 



(5) Add to each portion with a sterile 

 pipette 5.0 cc. of Merck's glycerol 

 pancreatin. 



(6) Keep one portion in the incubator 6 

 hours, another 24 hours and the other 

 48 hours. 



(7) Sterilize in streaming steam. 



(8) Neutralize with HCl. 



(9) Dilute each portion to 650.0 cc. and 

 add 2.0 g. NaCl to each portion. 



(10) Add glycerol and agar as usual 

 (amount not given). 



Sterilization: Not specified. 



Use: Cultivation of parasitic and sapro- 

 phytic forms. Authors reported that 

 some organisms developed better on the 

 medium with the longer pancreatin 

 digestion, others required a shorter 

 period. 



Variants: 



(a) Bosse cultivated diphtheria bacilli 

 on a medium prepared as follows: 



