VACCINE-THERAPY 23 



The culture in plate No. 1 will usually be too dense to 

 differentiate, but plates Nos. 2 and 3 can as a rule be used 

 for subculture. This is done by taking off a few colonies 

 with the platinum loop and inoculating a test-tube medium 

 and incubating them. Agar plates may be used in a 

 similar manner, but, owing to their rapidly - cooling 

 qualities, the work must be done very quickly. To grow 

 bacteria with the view of procuring a large supply of 

 bacterial emulsion, a convenient method is to pour the 

 medium into flat-sided bottles, allow it to set, and then 

 pour over the surface a small quantity of bacteria-laden 

 broth, taking care it is evenly diffused all over the face of 

 the medium, and that no surplus overflow exists. 



Anaerobic Cultivation. 



Many bacteria will not grow in the presence of oxygen 

 (free), and various methods are adopted to remove oxygen 

 from the atmosphere they grow in. Some of these 

 methods are very simple, others very complex. The simplest 

 are as follows : 



1. Take a narrow test-tube three-quarters filled with 

 sterile agar medium to which has been added a 10 per 

 cent, solution of glucose, place it in hot water, and boil 

 well for five or ten minutes. Cool and make solid by dip- 

 ping in cold water. The germ-laden needle (a large one) 

 should now be thrust into the axis of the medium, reaching 

 the bottom, rotated and withdrawn, and the sterile plug 

 inserted. The tube is then heated at the upper border of 

 the medium to seal the puncture, and a well-fitting rubber 

 cap applied. The tube is now ready for incubating. 



2. Another simple method is to insert a plug of cotton- 

 wool into a test-tube, place some pyrogallic acid on it and 

 moisten with caustic soda or potash, then a little layer of 

 cotton-wool, and fix on the mouth of the tube a tight-fitting 

 rubber cap. As the oxygen becomes absorbed, the cap 

 will be drawn in by suction. 



Four grammes of pyrogallic acid and 8 grammes of 



