PREPARATION OF MEDIA 35 



multiplication of other organisms. When required for use, a given 

 quantity is filtered through filter paper moistened with water (the 

 nitration takes some hours), heated up to the boiling point, and 

 made neutral to phenolphthalein with normal caustic soda solu- 

 tion. In the preparation of nutrient bouillon or other media, equal 

 quantities of this liquid and beef bouillon prepared as below are 

 mixed together. 



Preparation of Media 



The method of preparation will now be given, according to 

 recognised formulae, of the ordinary laboratory media likely to be 

 of service to the bacteriologist engaged in work upon milk, with 

 the addition of those peculiarly applicable to this special branch of 

 research. 



I. Nutrient beef bouillon — 



Take i pound of beef free from fat and connective tissue, 

 mince, add i litre of water, and allow to digest in a cool place 

 for eighteen hours, stirring occasionally. Strain through a fine 

 sieve and muslin, place the liquid in a clean enamelled saucepan, 

 heat up slowly to boiling point and allow to boil for five minutes. 

 Filter through damp paper into a clean flask. 



To make 2 litres of media, add to the litre of liquid thus 

 obtained i litre of the pig's stomach broth prepared as above, 

 and 10 grammes of common salt, and bring the liquid to the re- 

 quired reaction in the manner described under the head of neutral- 

 isation of media. Make up to 2 litres with distilled water, boil 

 for ten minutes,^ and filter through damp filter paper into flcisks 

 or tubes. Sterilise in the autoclave for fifteen minutes at 115°, or 

 in the Koch steriliser at 100° for thirty minutes on three successive 

 days. 



The bouillon can be cleared with white of q%^, but if the above 

 instructions are carefully carried out this will be found unnecessary, 

 and a perfectly limpid fluid will be the result. 



2. Glycerine bouillon — 



Add to bouillon prepared as above 6 per cent, of pure glycerine 

 immediately after the final filtration, and thoroughly incorporate 

 by stirring or shaking. 



^ If final sterilisation is to take place in the autoclave the bouillon should, in 

 order to get rid of the excess of earthy phosphates which may subsequently 

 trouble it, be autoclaved at this stage for fifteen minutes at 115°. 



