524 PATHOGENIC MICROORGANISMS 



.employed. Amoss has found that young rabbits are more satisfac- 

 tory than older ones for this purpose, and he uses rabbits weighing 

 between 1500 and 1800 grams. He grows his meningococci on glu- 

 cose agar slants, and washes up the growth in salt solution. 0.001 

 of a culture is inoculated as the first dose. For the rapid production 

 of agglutinating sera, he injects his rabbits for three succeeding 

 days, giving a rest of five days, and then another course of three 

 days injection. He bleeds the animal two or three days after the 

 second course of inoculation. For ordinary purposes, the slow 

 method of three or four day intervals, about five or six injections, 

 with bleeding eight or nine days after the last injection, may be 

 used. Among English workers, Hine 29 injects culture suspensions 

 grown on 25 per cent hemoglobin serum agar, killed at 65 and 

 brought to a standard opacity. 0.5 per cent carbolic acid is added 

 for preservation. He standardizes all his suspensions by opacity 

 comparisons against suspensions of freshly precipitated barium 

 sulphate. He compares by diluting his suspension in a test tube of 

 similar dimensions as the standard tube, until the image of a small 

 flame is just visible in the same distance from the flame as in the 

 case of the standard tube. With such suspensions he immunizes rab- 

 bits, beginning with an injection of two doses of five hundred 

 million cocci at an interval of one hour. Six days later he gives three 

 million cocci, and, if the serum is satisfactory on the eighth day 

 later, he bleeds. This method was satisfactory in the hands of Hine, 

 with types I and III. With the other types he has had to give larger 

 and more frequently repeated doses. In all such immunization, 

 experience and judgment, with frequent titration of samples of the 

 rabbit serum taken from the ear, are necessary. 



Nicolle 30 at the Pasteur Institute uses for immunization, pow- 

 dered meningococcus antigen prepared from growth on agar slants 

 by suspension in salt solution, centrifugation and drying. Hine also 

 recommends the use of rabbits ranging from 800 to 1500 grams. 



Agglutination Technique ivith Meningococci. Agglutination of 

 meningococci present considerable difficulties because of the relative 

 inagglut inability of many meningococcus cultures. This is a peculiar- 

 ity of these organisms which has necessitated much investigation and 

 many technical modifications. Hine has found that allowing the 



29 Hinc, Med. lies. Committee, Spec. Eep., Series 3, No. 3, p. 99, 



30 Nicolle, 



