CULTUEE MEDIA FOR CULTIVATION OF MICROORGANISMS 



573 



1837. Paneth's Glucose Ascitic Fluid Agar 



Constituents : 



1. Distilled water 3000.0 cc. 



2. Liebig's meat extract 12.0 g. 



3. NaCl 15.0 g. 



4. Peptone (Witte's) 30.0 g. 



5. N/IH2SO4 17.5 cc. 



6. Agar 85.0 g. 



7. Glucose 50.0 g. 



8. Ascitic fluid 

 Preparation : 



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



(2) Add 6 to (1). 



(3) Allow to stand for several hours. 



(4) Boil in the steamer for two hours. 



(5) Filter thru gauze. 



(6) Mix the whites of eight eggs with 

 150.0 cc. distilled water. 



(7) Cool the liquid agar to 50°, add (6) 

 and mix well. 



(8) Pour into flasks and heat at 100° in 

 the steamer for two hours. 



(9) Filter thru wet cotton. Gauze is 

 placed on top of the cotton. 



(10) Adjust the reaction from 1.8 to 2.2. 



(11) Add 2.0% glucose (about 50.0 g.). 



(12) Heat for 20 minutes in a steamer. 



(13) Distribute into large test tubes in 

 about 15.0 to 20.0 cc. lots. 



(14) Distribute sterile ascitic fluid into 

 large sterile test tubes. 



(15) Liquify the agar and cool to 40 to 

 42°C. 



(16) Draw, under aseptic conditions, 

 about 2.0 cc. of infected blood di- 

 rectly into the liquefied agar and 

 add one tube of ascitic fluid (amount 

 not specified) to the agar. Mix. 

 (In the work of Plotz, Olitsky and 

 Baehr, 2.0 cc. of ascitic fluid is 

 added.) 



(17) After cooling cover with plain agar 

 or paraffin. 



Sterilization: Sterilize (13) for 20 minutes 

 in the autoclave at 10.0 kilograms pres- 

 sure or in the steamer on two or better 

 four successive days for 20 minutes each 

 day. 



Use: Cultivation of Bacterium typhi 

 exanthematis. 



Reference: Paneth (1916 p. 647). 



1838. Plotz, Olitsky and Baehr's Ascitic 

 Fluid Agar 



Constituents : 



1. Glucose agar (2.0%) 1000.0 cc. 



2. Ascitic fluid 200.0 cc. 



3. Plain agar 

 Preparation : 



(1) Prepare nutrient agar using Liebig's 

 meat extract and containing 2.0% 

 glucose and 2.0% agar. 



(2) Distribute in 20.0 cc. lots in tubes 

 15.0 cm. X 2 cm. 



(3) Cool the sterile melted agar tubes to 

 40°C. and add to each tube 4.0 cc. 

 ascitic fluid and 2.0 cc. of infected 

 blood. 



(4) Mix by pouring back and forth into 

 sterile test tube, avoiding air bubbles. 



(5) When the agar has solidified add 

 sterile plain agar to a depth of 2.0 

 cm. to each tube. 



(6) Incubate at 37.5°C. 

 Sterilization: Method not given. 



Use: Cultivation of B. typhi exanthematici 

 (supposed cause of typhus fever), from 

 blood. 



Variants : Baehr and Plotz gave the follow- 

 ing method of preparation: 



(1) Mix 12.0 g. Liebig's meat extract, 

 15.0 g. NaCl, 30.0 g. Witte's peptone, 

 17.5 g. N/1 H2SO4, and 85.0 g. agar 

 shreds in 3000.0 cc. water. 



(2) Place in an Arnold steam sterilizer 

 for two hours. 



(3) Filter rapidly thru several layers of 

 gauze to remove water. 



(4) Mix the whites of eight eggs with 

 150.0 cc. distilled water. 



(5) Cool (3). 



(6) Mix (5) and (4) and shake thoroly. 



(7) Place in Arnold sterilizer (100°C.) 

 for two hours. 



(8) Decant clear agar and filter thru 

 moistened absorbent cotton. 



(9) Adjust to acidity of 0.9 to 1.1 to 

 phenolphthalein. 



(10) Add 2.0% glucose to (9). 



(11) Distribute into 2 x 20 cm. test tubes 

 in about 20.0 cc. lots (half full). 



(12) Sterilize (11) at 10.0 kg. pressure for 

 20 minutes or in an Arnold on 3 or 

 4 successive days for 20 minutes. 



