CULTURE MEDIA FOR CULTIVATION OF MICROORGANISMS 



327 



(2) Mix 500.0 g. beef heart or human 

 placenta with 1000.0 cc. water. 



(3) Make faintly alkaline to litmus 

 using N KOH or N Na-.COs. 



(4) Heat slowly to 70-80° for 5-10 

 minutes. 



(5) Cool to 37°C. and add 1.0% pan- 

 creatic extract or "Bacto" trypsin. 



(6) Incubate at 37°C. for 2-5 hours. 



(7) Control the progress of digestion by 

 repeated Biuret and tryptophane 

 tests. In case it is necessary to 

 extend the digestion over a period of 

 6 hours add chloroform or toluene. 



(8) When the process is sufficiently ad- 

 vanced, render slightly acid with 

 glacial acetic acid and boil slowly 

 for 15 minutes. 



(9) Either filter or decant the clear 

 fluid which results on placing the 

 digest over night in a cool place. 



(10) Adjust the reaction as desired. 



(11) Dissolve 2, 3, 4, 5 and 8 in (10). 



(12) Heat for 15-30 minutes in the 

 steamer at 100 °C. 



(13) Filter again if necessary. 

 Sterilization: Sterilize at 100°C. on 3 con- 

 secutive days if not to be used at once. 



Use: General inexpensive culture medium. 

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



1137. Stickel and Meyer's Digest Solution 



Constituents : 



1. Water, tap 4000.0 cc. 



2. Liver, placenta or blood 



clot 400.0 g. 



3. Stomach, hog 400.0 g. 



4. K2HPO4 8.0 g. 



Preparation : 



(1) Wash clean and mince fine 5 or more 

 large pig's stomachs. 



(2) Mince an equal amount of clean 

 pig's or beef liver, cheap fat free 

 beef, placenta or blood clots. 



(3) Mix 400.0 g. of (1), 400.0 g. of one 

 of (2) and 40.0 g. of Baker Chemical 

 Co. HCl in 1000.0 cc. of tap water 

 at 50 °C. 



(4) Incubate at 50°C. for 18 to 24 hours. 



(5) Make a Biuret and tryptophane test. 

 When both are + the digest is yel- 

 lowish green and contains very 

 little undigested debris. 



(6) Transfer to large bottles and steam 

 for 10 minutes at 100°C. to stop 

 digestion. 



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

 line to litmus using 2N KOH or 

 2 normal Na2C03. 



(8) Cool to 37°C. and add 1.0% pan- 

 creatic extract or "Bacto" trypsin. 



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

 10 hours depending on the reaction 

 of the trypsin and the digestion 

 desired. Control the process by 

 repeated tests for tryptophane. 



(10) When trypsinizing is sufficiently ad- 

 vanced render reaction slightly acid 

 <vith glacial acetic acid, and bring 

 slowly to boiling point for 10 

 ininutes. 



(11) Filter thru paper or keep in cool 

 place over night and decant the 

 clear liquid in the morning. 



(12) Add K2HPO4 and adjust the reaction 

 faintly alkaline or to the desired 

 H-ion concentration. 



(13) Heat in steamer at 100° for 15 

 minutes. 



(14) Correct reaction and filter thru 

 paper. 



(15) Distribute in receptacles used for 

 culture. 



Sterilization: Sterilize at 100°C. for 30 min- 

 utes on 2 successive days or at 10 pounds 

 pressure for 15 minutes. 



Use: General inexpensive culture medium. 



Variants: The authors prepared a medium 

 as indicated above steps (1) thru (7). 

 The remainder of the preparation was as 

 follows: 



(8) Cool to 37°C add 1.0% pancreatic 

 extract or 40.0 g. trypsin, 8.0 g. 

 K2HPO4, 4.0 g. CaCOs, and 1.0% of a 

 24 hour old broth culture of B. sac- 

 char olyte. 



(9) Incubate at 37°C. for 12-18 hours and 

 control the digestion by tryptophane 

 tests and the removal of carbohy- 

 drates by the gas formation in fer- 

 mentation tubes. 



(10) When the digest is sugar free steam 

 15 minutes. 



(11) Adjust to desired reaction and steam 

 another 15 minutes. 



(12) Filter thru paper. 



(13) Distribute as desired. 



