202 MICROSCOPIC AND CULTURAL STUDY OF BACTERIA 



color from the acid colonies. For rapid isolations the medium with 

 the normal percentage of agar is preferable. 1 



The Technique of Inoculation of Endomedia is described on page 231. 



Lactose Litmus Agar. I. Prepare 1 per cent, lactose nutrient agar 

 by adding 10 grams of C. P. lactose (free from dextrose) to one liter 

 of plain nutrient agar. Adjust the reaction to a point slightly alkaline 

 to litmus. Tube and sterilize in the Arnold sterilizer. 



II. Prepare an aqueous solution of litmus either a 5 per cent, 

 solution of purified litmus (Merck) or a 1 per cent, solution of azo- 

 litmin (Kahlbaum) and sterilize. 



To Use Lactose Litmus -Agar. Add about 1 c.c. of sterile litmus 

 solution to a sterile Petri dish and pour over it the melted lactose 

 agar, previously inoculated with the desired material. For water and 

 milk, add 1 c.c. of water or milk (diluted to the proper degree) to the 

 Petri dish before adding the lactose agar. Mix intimately by rotating 

 gently, allow to harden, and incubate. 



Those bacteria which ferment lactose with the production of acid 

 appear as red colonies. Non-lactose-fermenting organisms appear as 

 blue colonies. 



Blood Agar. Blood is drawn with aseptic precautions from the 

 carotid or femoral artery of a dog or rabbit into a sterile flask con- 

 taining beads. The blood is defibrinated by prolonged agitation and 

 added to plain (not dextrose) nutrient agar previously melted and 

 cooled to 45 C., in the proportion of 2 c.c. of blood to 10 c.c. of agar. 

 Small amounts of blood may be withdrawn directly from the heart 

 of an animal without difficulty, provided a small hypodermic needle 

 is used. The blood may be injected directly into the melted agar 

 without defibrination. 



Occasionally human blood is added to agar; if a series of agar slants 

 are prepared it is possible to convert them into blood agar with a small 

 amount of blood, as follows : 



Withdraw 10 c.c. of blood, using aseptic precautions, from the 

 median basilic vein, in a large syringe. Inject the blood at once into 

 four times the volume of plain nutrient agar melted and cooled to 45 

 C. Mix at once and run 2 c.c. over the slanted surface of each agar 

 slant, and allow to harden in the inclined position in such a manner 

 that a uniform layer of the blood-agar mixture is obtained: Incubate 

 to prove sterility. 



1 Kendall and Day, Jour. Med. Res., 1911, xxv, 95. 



