BACTERIAL INOCULATION 221 



of accuracy and the proportionate number of bacteria 

 to the red blood-cells counted, determined as follows : 

 If in fields containing 1000 red blood-cells 500 bac- 

 teria are counted we have the proportion 



1000 : 500 :: 5,000,000,000 : x. 



X =z 2,500,000,000, or the number of bacteria per c.c. of suspension. 



This suspension is obviously too concentrated and 

 may be conveniently diluted as follows: If it is de- 

 sired to prepare the bacterin in 10 c.c. containers, so 

 that each cubic centimetre shall contain 400,000,000 

 bacteria, we have the formula 



400,000,000 X 10 



2,500,000,000 



=^ x = 1.6 c.c. 



Therefore to 1.6 c.c. of the original suspension 

 must be added 8.4 c.c. of the diluting fluid, phenol-salt 

 solution. The bacterin is then sterilized by submer- 

 sion in a water-bath for one-half to one hour at the 

 thermic death-point of the particular bacterium, usu- 

 ally about 60° C. After removal from the bath, a 

 drop of the bacterin is cultured for a day or two in 

 order to test its sterility before employment thera- 

 peutically. Other methods of sterilization occasion- 

 ally utilized include chemical, cold and maceration by 

 prolonged shaking. 



Preparation of Bacterins by Autolysis. — Certain 

 investigators have attempted to improve bacterins 

 and enhance their immunizing properties by subject- 



