CULTURE MEDIA FOR CULTIVATION OF MICROORGANISMS 



519 



(6) Add 1.0 cc. of a 1 to 1000 aqueous 

 solution of brilliant green and 

 1.0 cc. of a 1.0% solution of picric 

 acid to each flask. 



(7) Pour into large petri dishes. 

 References: Bezan<;on (1920 p. 345), Har- 

 vey (1921-22 p. 91), Stitt (1923 p. 49). 



1708. Klinger's Malachite Green Extract 

 Agar 



Constituents : 



1. Water 2000.0 cc. 



2. Meat extract (Liebig's) 20.0 g. 



3. Peptone (Witte) 20.0 g. 



4.NaCl 10-Og- 



5. Agar 80 g- 



6. Malachite green (Hochst 120) 

 Preparation : 



(1) Add 2, 3, 4 and 5 to 1. 



(2) Heat for either 3 hours in streaming 

 steam, or 2 hours at 110° or 1 hour 

 at 120°C. 



(3) Filter thru a thick layer of cotton. 



(4) Remove 50.0 cc. of the agar and 

 measure the rest. 



(5) Allow the agar to solidify. 



(6) After solidification add 0.05 g. Hochst 

 120 malachite green to each 100.0 cc. 

 of agar. 



(7) Liquefy the agar. 



(8) Determine the reaction. of the 50.0 cc. 

 of agar removed in (4) and add suffi- 

 cient normal NaOH to melted mala- 

 chite green agar so that the alkalinity 

 for 100.0 cc. of agar is 1.0 cc. of normal 

 NaOH using phenolphthalein as an 

 indicator. 



Example: If 0.9 normal cc. are re- 

 quired to neutralize the 50,0 cc. agar, 

 then 1.8 cc. would be required for 

 100.0 cc. H one wishes to adjust the 

 reaction of 400.0 cc. of malachite 

 green agar add 4 X 0.8 = 3.2 cc. 

 normal NaOH. 

 (9) Pour into sterile plates. 

 Sterilization: Not specified. 

 Use: Isolation of typhoid bacilli. The 

 author reported that other forms were 

 inhibited. 

 Variants: Zipfel regenerated malachite 

 green agar in the following manner: 

 (1) Remove the malachite green agar 

 slants and plates from tubes and 

 Petri dishes. 



(2) Add 3.0% HCl to used malachite 

 green agar slants or plates until the 

 HCl covers the agar. Stir. 



(3) Allow the acid to react for one hour, 

 stirring continually. 



(4) Pour the agar on a sieve. 



(5) Wash the pieces of agar with a stream 

 of water until the wash water is 

 clear. 



(6) Soak the agar in water for 24 hours, 

 changing the water often. 



(7) Place the agar on a sieve at the end 

 of this time and allow to drip free 

 from water. 



(8) Liquify the agar particles in stream- 

 ing steam. 



(9) To a liter of (8) add 8.0 to 10.0 cc. of 

 a 10 0% soda solution, 4.0 cc. of a 

 filtered and sterilized solution of 

 20,0%, peptone and 20.0% meat 

 extract or meat equivalent. 



(10) Sterilize for one hour in the steamer. 



(11) The reaction of the agar determines 

 the amount of malachite green to be 

 added. There is an optimum for 

 each batch of agar, and must be 

 determined in each case. 



References: Klinger (1906 p. 52), Zipfel 

 (1917-18 p. 479). 



1709. Kohler's Basal Agar 



Constituents : 



1. Nutrient agar 1000.0 cc. 



Preparation: (1) Dissolve one of the added 



nutrients in nutrient agar. 

 Sterilization: Not specified. 

 Use: To study the effect of acids and other 

 materials on the growth of the typhoid 

 bacillus. Other investigators used sim- 

 ilar media for a variety of purposes. 

 Added nutrients: The author added 0.05, 

 0.1, 0.15, 2, 0.25, 0.3, 0.35, 0.4, 45, 0.5, 

 0.55, 0.6, 0.65, 0.7, 0.75, 0.8, 0.85, 0.9, 95, 

 1.0, 1.5 or 1.1% of one of the following 

 materials: 



Lactic acid HNO3 (30%) 



Citric acid H2SO4 (97%) 



H3PO4 (2.0%) NaOH (33%) 



Tartaric acid KOH (33%) 



Alum Methyl violet 



Carbolic acid Fuchsin 



HCl (25%) 

 Variants : 



(a) Thoinot added any desired amount of 



