12 LABORATORY BACTERIOLOGY 



Put 5 cc. in each of 10 small sterile test tubes. 



Put 10 cc. in each of 12 large sterile test tubes. 



Put the remainder in the flask which contained the bouillon. 



17. The preparation of nutrient gelatin. Take a flask of 

 bouillon containing 300 cc. and pour it into a small agate iron 

 dish, add 30 grams of sheet gelatin, and heat, with frequent 

 stirring, in a water bath, until the gelatin is dissolved. Allow 

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

 add the white of one egg and mix it thoroughly by stirring, 

 or, better, by pouring the gelatin many times from one flask 

 or beaker to another. After the egg albumen is completely 

 diffused, return the liquid gelatin to the large covered water 

 bath and boil until the egg albumen is firmly coagulated. 

 This takes about 20 minutes. It is now ready for filtering, 

 which must be done while the gelatin is hot. Filter through 

 properly folded l but ordinary filter paper, first moistened with 

 boiling water. Distribute the filtrate as directed. In pouring 

 the gelatin into the tube use a small 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 

 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 the other 

 tubes with a moist cloth, label, and place them in the locker, 

 where they can be kept until used. 



18. The preparation of 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, add 75 cc. 

 of distilled water, and boil over a gas flame, with constant 



1 For illustrations and directions for folding filter paper, see Abbott's 

 Principles of Bacteriology, 6th edition, p. 101. Filter paper already 

 folded may be procured. 



