[ 78? ] 



(5) When a pure culture of the virulent virus has been 

 obtained, a portion of it, say one flask out of three, is left for 

 ten days in the etuve, and the other two drawn out. The 

 one remaining in the etuve for ten days gives rise to a sporu- 

 lar virulent vaccine, which is useful for testing whether vac- 

 cination has been done or not. In other words, when animals 

 have been vaccinated with the premier vaccine, and twelve 

 days after with the deuxieme vaccine, a portion ot them 

 ought to be vaccinated twelve days after with the virulent 

 vaccine also, to see if it kills the animals, or the previous 

 vaccinations have had the effect of protecting them from the 

 effects of the virulent vaccine. If the animals die something 

 must have been wrong with the preparation of the vaccine or 

 the inoculation, and the operation has to be gone through 

 again. It is thus necessary to have a stock of virulent vac- 

 cine also. When the virulent virus has been ten days in the 

 etuve it is taken out and sucked into sterilized pipettes, the 

 two ends sealed, and the sporular virulent vaccine " seed" is 

 thus preserved for more than a year, as the spores of bacillus 

 anthracis can live without oxygen. 



(6) The premier and deuxieme vaccines are also kept in 

 stock in sealed tubes in their sporular state. When the 

 virulent vaccine is taken out of the stove on the third day 

 in its vegetating or filamentous condition it is subjected 

 to the attenuating process, i. ., it is kept in the Etuve 

 Arsonval for 12 days at 42 C. or 43 C. to prepare the 

 deuxieme vaccine, and for 20 days to prepare the premier 

 vaccine, which is more attenuated and weaker. This high 

 temperature attenuates the vaccine. It is then taken out of 

 the stove, and left for 10 days in the Etuve Pasteur for the 

 vaccines to change into the sporular state. When it is in the 

 Etuve Arsonval for 12 days or 20 days for attenuation it re- 

 tains its filamentous character ; but when brought back again 

 to the temperature of 28 C. to 32 C. in the Etuve Pasteur, 

 the most favourable temperature for culture, the vegetative 



