General Bacteriology. 



be so arranged that a temperature of 150 C. can be quickly reached and readily main- 

 tained. In such a sterilizer all glassware to be used for the reception of culture media, 

 such as flasks, test-tubes, Petri dishes, etc., is submitted to a temperature of 140-150 C. 

 for 1 hour, or until the cotton plugs are slightly browned; this change being due to the 

 incipient charring of the cotton. The test-tubes are placed erect in square baskets made 

 of galvanized iron wire. When the air in the sterilizer has cooled to about 40 C. the 

 glassware can be taken out and stored ready for use. The Petri dishes are not to be opened 

 until used for culture purposes. 



REFERENCES. A. 71 and 121; H. 32; L. & K. 74; M. & R. 36; N. 159; McF. 

 106; P. 223; S. 51. 



SPECIAL DIRECTIONS. All glassware prepared in I. is to be sterilized for one 

 hour at 150 C. The small pipettes should be placed in brass tubes, provided for the 

 purpose, and also sterilized. 



EXERCISE IV. PREPARATION OF BOUILLON. 



GENERAL DIRECTIONS. Any one of the three methods (A B or C) may be used. 

 They are arranged in order of preference, but method C is the most convenient, and 

 hence most used. 



A. 



B. 



Secure meat as 

 under A a, add 1 liter 

 of distilled water, 

 weigh (see e below), 

 cook for ^ hour at 

 about 70 C. , and pro- 

 ceed as directed under 

 e below. 



Weigh out three 

 grams of beef extract 

 (such as Liebig's), 

 add 1 liter of water, 

 and then proceed as 

 directed under e be- 

 low. 



a. From 500 grams (1^-lb.) of lean 

 beef, remove the fat and connective tis- 

 sue and mince (Hamburg steak). 



6 . Add 1 liter of distilled water and 

 after thoroughly shaking set in ice chest 

 for 12 to 24 hours. 



c. Squeeze through a cloth and add 

 enough distilled water to make 1 liter 

 and place in vessel to cook. This may be 

 done either in a flask which is heated in a 

 water-bath or a sterilizer, or in a rice 

 cooker. In this case use a 50 % solution of 

 calcium chloride in outer vessel instead 

 of water as by this means the contents 

 of the inner vessel can be brought to a 

 rapid ebullition, something impossible by 

 the use of water alone. 



d. Boil \ hour and make up loss of 

 water. 



e. Add to any of the above solutions: 



1% (10 gms.) peptone (Witte) and -5-% (5 gms.) common salt (NaCl), then weigh 

 solution, with vessel, so that the water which is subsequently driven off in cooking can 

 be accurately replaced. 



/. Heat until ingredients are in solution, then restore the water lost by evaporation. 



g. Neutralize or render slightly alkaline. This is a very important step and calls for 

 great care. Method A is more accurate and should be employed for special or research 

 work. For ordinary routine work B may be employed. 



C. 



