II] PREPARATION OF CULTURE MATERIAL 35 



can be easily driven off by placing the test-tubes in a 

 slanting position in the hot incubator (37°) and leaving 

 them here for a few days. Owing to the large surface of 

 evaporation the condensation water is soon got rid of. 



5. KanthacKs serum is a mixture of solidified ascites 

 fluid and Agar, which far surpasses all other media for the 

 isolation of the diphtheria bacilli, even when in a given 

 material these latter are almost swamped by other microbes : 

 the cultivation of this material rubbed over the slanting 

 surface of this serum (solidified) will at twenty-four to forty- 

 eight hours' incubation show and pick out in a remarkable 

 manner the colonies of the diphtheria bacilli in almost pure 

 culture. It is prepared thus : Ascites fluid is received by 

 sterile trocar and tubing into a sterile flask plugged with 

 sterile cotton-wool. For each 100 cc. of ascitic fluid to be 

 treated, take 2 grams of Agar-Agar and treat as follows : 

 2 to 3 cc. glacial acetic acid in 500 cc. of distilled water ; 

 put Agar in this solution and allow to soak for thirty 

 minutes, wash in several lots of tap-water and finish with 

 distilled water, thoroughly drain. For each 100 cc. of 

 ascitic fluid add 2 cc. of a 10 per cent, solution of caustic 

 potash, and very thoroughly mix. Add this to the Agar, 

 now add 6 per cent, of glycerine, and place in steamer and 

 steam at 100° C. for one and a half hours; filter through 

 Chardin's filter paper, decant into tubes and steam at 

 100° C. on three successive days for thirty minutes. 



6. Nutrietit Agar-Agar. — " Nutrient Agar " is Japan 

 isinglass. This was first used for preparation of solid 

 culture medium by Koch. The best and quickest mode of 

 preparing nutrient Agar is the following modification of 

 Tischutkin's method described in Schenk's Elements of 

 Bacteriology, English translation, p. 44. It is the nutrient 

 Agar which I now use, being easily and quickly pre- 



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