90 LABORATORY EXERCISES IN BACTERIOLOGY. 



to dissolve. When the gelatine has been dissolved, cool the preparation 

 to 60 C. or less, and add thereto one-half the white of an egg dissolved 

 in twenty-five cubic centimeters of boiled water (ten per cent, solution 

 of commercial egg-albumen may be substituted), mixing it well with 

 the liquid by stirring. Boil for ten or fifteen minutes to coagulate the 

 egg-albumen and to reduce the volume by evaporation to the original 

 amount or less. Filter hot through a moistened filter paper which has 

 been folded after the manner customary in the chemical laboratory, to 

 separate the coagulated egg-albumen (albumen added and coagulated 

 in order that solid particles which might interfere with the transparency 

 of the product may be caught in the meshes of the coagulum and thus 

 be removed) ; correct volume (correct to original weight, i. e,, weight of 

 bouillon plus weight of gelatine). While yet hot determine reaction and 

 adjust to standard. If medium becomes clouded, again boil and filter. 

 Distribute half to tubes, and sterilize by fractional steaming; retain re- 

 mainder in small stock-flask and sterilize in the same manner. 



3. Peptone Agar-agar ("Agar"). This medium is made by the addition of 

 two per cent, of agar-agar (a vegetable gelatine derived from an alga found on the 

 Eastern Asiatic coast) to bouillon. The purpose of the addition of this substance 

 is the same as that of the addition of gelatine i. e., the solidification of the mass; 

 but as the medium after preparation is not liquefied by temperatures below 85 to 

 90 C. (again setting at about 40 C.), it is better suited than gelatine for culture of 

 organisms requiring incubation. In the manufacture it is necessary that the agar 

 should have been thoroughly dissolved into a limpid liquid for ready filtration ; 

 this is accomplished by prolonged boiling (one or two hours), its property of con- 

 gelation not being interfered with by such exposure to heat. The presence of excess 

 of acid in the material to which the agar is added is more or less harmful to its solidi- 

 fying power, for which reason the bouillon should have been neutralized (litmus re- 

 action sufficient) before the agar is added. The product is not quite so clear as gelatine 

 at best. This medium is not affected by the proteolytic ferments. 



Exercise 28. Five grams of finely chopped or ground agar are boiled 

 for one or two hours in one hundred cubic centimeters of water in a beaker 

 or other vessel, water being added from time to time to prevent drying. 

 To this is added at the close of boiling 250 cubic centimeters of the bouillon 

 prepared in exercise 25 (or same amount of bouillon freshly prepared 

 up to the stage of sterilization, if stock bouillon be not on hand) ; and 

 the mixture boiled until evaporation has reduced it approximately to 

 the desired weight (weight of bouillon plus five grams of agar). It is 

 now cooled to 60 C., or less, and one-half the white of an egg dissolved 

 in twenty-five cubic centimeters of boiled water added and stirred into 

 the preparation. Reboil for ten or fifteen minutes to coagulate the egg- 

 albumen and filter through a folded and moistened filter paper. If it is 



