192 Cultivation of Micro-organisms 



standard reaction of media, but with the recommendation that the optimum 

 growth reaction be always recorded with the species." 



BOUILLON OR BROTH 



This is one of the most useful and most simple media. It can be 

 prepared from meat or from meat extract, and is the basis of most 

 of the culture-media. The addition of 10 per cent, of gelatin makes 

 it "gelatin;" that of i per cent, of agar-agar makes it "agar- 

 agar." 



I. To Prepare Bouillon from Fresh Meat. — To 500 grams of finely 

 chopped lean, boneless beef, 1000 c.c. of clean water are added and 

 allowed to stand for about twelve hours on ice. At the end of this 

 time the liquor is decanted, that remaining on the meat expressed 

 through a cloth, and then, as the entire quantity is seldom regained, 

 enough water added to bring the total amount up to 1000 cc. This 

 liquid is called the meat-infusion. To it 10 grams of Witte's or 

 Fairchild's dried beef-peptone or "Bacto-Peptone" made by the 

 Digestive Ferments Co., Detroit, Michigan, and 5 grams of sodium 

 chlorid are added, and the whole boiled until the albumins of the 

 meat-infusion coagulate, titrated or otherwise corrected for acidity, 

 boiled again for a short time, and then filtered through a fine filter 

 paper. It should be slightly yellow and perfectly clear and Umpid. 

 Smith,* referring to bouillon intended for the culture of diphtheria 

 bacilli for toxin, says that when the peptones are added before boiling 

 most of them are lost, and therefore recommends that the meat- 

 infusion be boiled and filtered and the sohd ingredients added and 

 dissolved subsequently. The reaction, which is strongly acid, is 

 then carefully corrected by titration according to the directions 

 already given. 



The filtered fluid is dispensed in previously sterihzed tubes with 

 cotton plugs — about 10 cc. to each — or in flasks, and is then sterilized 

 by steam three successive days for fifteen to twenty minutes each, 

 according to the directions already given for intermittent steriliza- 

 tion, or superheated in the autoclave. 



The loss of water during boiling is an important matter to bear 

 in mind, as unless properly replaced it is the cause of disproportion 

 between the fluids and soHds of the media. The quantity must 

 therefore be measured before filtration and enough water added to 

 replace what has been lost. Measuring before filtration is compara- 

 tively easy with bouillon, but difiicult with heavy liquids, like the 

 gelatin and agar-agar solutions. To overcome this difficulty it is 

 best to make the entire preparation by weight and not by volume. 

 A pair of platform scales with shding indicators will first balance 

 the empty kettle and then show the correct quantity of each added 

 ingredient. After boiling, the kettle can be returned to the" scale 

 and the exact quantity of water to be added determined. 



* "Trans. Assoc. Amer. Phys.," 1896. 



