19 



beaker or graduate and see that the gelatin does not touch 

 the sides of the upper part of the tube. Stand the tubes in 

 a wire basket and sterilize them by boiling in a closed water 

 bath or by steaming in the Arnold's steam sterilizer for 30 

 minutes. The small flasks can be sterilized in the same 

 manner. Place tubes and small flasks in the incubator and 

 allow them to remain there for two days. If the gelatin in 

 any of the tubes becomes cloudy the medium in those tubes 

 must be rejected. Carefully wipe all of the other tubes with 

 a moist cloth, label and place them in the locker where they 

 can be kept until used. 



1 6. Nutrient agar. Weigh out 3 grams of agar and cut 

 it into small pieces with a pair of scissors. Put the finely 

 cut agar into an agate iron dish and add 50 c.c. of distilled 

 water and boil it over a gas flame with constant stirring, to 

 prevent scorching, until the agar is dissolved, giving a thick 

 homogeneous pasty substance. Pour 300 c.c. of bouillon 

 ( 10) from a flask into an agate iron cup and to it add the 

 dissolved agar. Place the dish containing the mixed agar 

 and bouillon in a closed water bath and boil for 20 minutes. 

 Then cool it to a temperature between 45 and 50 C. and 

 add the white of one egg and thoroughly mix it in the 

 liquid agar. This is easily accomplished by pouring it from 

 one beaker to another. When the egg albumen is dissolved 

 the agar is returned to the water bath and boiled vigorously 

 until the white of the egg is coagulated. This usually takes 

 about 20 minutes. Filter the agar immediately, while hot, 

 through ordinary filter paper which has been moistened with 

 boiling water. Distribute the filtrate in small and large 

 tubes, as directed. Sterilize, label, and store the agar in the 

 same manner as the gelatin. 



