BACTERIOLYTIC REACTIONS 269 



grams) of an eighteen-hour-old culture of the suspected organism, sus- 

 pended in 1 c.c. of broth. A second animal ( B) is given ten titer doses 

 ( = 2 milligrams) with the same quantity of organisms. A third (C) 

 receives 50 multiples of the titer dose, i. e., 10 milligrams, of normal 

 serum, however, but taken from an animal of the same species as 

 that furnishing the immune serum, together with the same quantity 

 of organisms as A and B, while a fourth guinea-pig ( D) is injected 

 with the same dose of bacteria, but without any serum. The animals 

 should all be of about the same weight (250 grams), and are all 

 injected intraperitoneally. To this end it is recommended to make 

 a small incision through the skin and to inject through a cannula 

 with a blunt point. By the aid of glass capillaries a droplet of 

 the peritoneal fluid is then procured through the same incision, 

 immediately after the injection, a second one twenty minutes later, 

 and a third one at the expiration of one hour. The specimens are 

 examined as hanging drops with an oil immersion lens. If the 

 organism under consideration is the cholera vibrio, typical granule 

 formation and lysis will be observed in specimens A and B after 

 twenty minutes already, and at the latest at the expiration of one hour; 

 while in C and D there will be large numbers of actively motile 

 organisms or such at least in which the form has been well preserved, 

 the C animal being the control to A and B. The object of injecting 

 D is merely to prove that the organism in question is virulent and 

 this animal as well as C, of course, should die, while A and B remain 

 alive. If the result then turns out as just indicated, the inference is 

 justifiable that the organism under examination was really the 

 cholera vibrio. 



2. Pfeiffer's Test as Applied to the Recognition of Recent Cholera 

 Infections. In this case the individual's serum is diluted with broth 

 in the proportion of 1 to 20, 1 to 100 and 1 to 500, when guinea-pigs are 

 each inoculated as described above with 1 c.c. of various dilutions, 

 together with one oese ( = 2 milligrams) of an eighteen-hour-old agar 

 culture of a virulent cholera strain. If extensive bacteriolysis can then 

 be demonstrated at the expiration of twenty minutes, or at most, 

 an hour, the inference is justifiable that the person has recently 

 passed through an attack of cholera. 



PREPARATION OF THE CHOLERA IMMUNE SERUM. The cholera 

 immune serum which is required in test 1 (above) is prepared as 

 follows: A number of rabbits are each injected intraperitoneally 



