PRODUCTION OF AGGLUTINATING SERA I Op 



i :io respectively. Each cover slip is inverted over a hollow slide pro- 

 tected by vaseline, and examined microscopically. A control with normal 

 serum and also one with culture alone should be made. 



For the identification of bacteria only highly agglutinating 

 Production animal sera can be employed. Rabbits, goats and horses, 

 of Aggluti- are most suitable for such experiments. The best results 

 nating Sera. are obtained when the animals are immunized intravenously 

 by repeated injections with gradually increasing doses of dead 

 bacteria (killed at 60 C.). Usually two to three injections of 1/4 to i agar 

 culture of bacteria suspended in saline solution suffice to give an agglutinat- 

 ing titer of i to 5000. The serum should be withdrawn eight to ten days 

 after the last injection. With typhoid bacteria one may attain a strong 

 agglutinating serum in a rabbit by the intraperitoneal injection of i c.cm. 

 of a twenty-four hours' live broth culture. This is to be repeated in 7 to 10 

 days. As a matter of course, the titer of the serum should be tested from 

 time to time, because the height of the antibody curve can only reach a 

 certain point. When a sufficient strength is obtained the animal is bled. It 

 is not possible to produce equally strong agglutinating sera for all bacteria. 

 The agglutinins belong to the class of the more resistant serum sub- 

 stances. By the addition of one drop of pure carbolic acid they can be 

 preserved on ice for a long time. Heating variously affects the different 

 bacterial agglutinins. The agglutinins for pest and tubercle bacilli are 

 destroyed at 56 C. while other bacteria are not influenced by even higher 

 temperatures. The animal from which the agglutinating serum has been 

 obtained also influences to a great degree the resistance toward heat. 

 Thus the typhoid agglutinating serum derived from the horse is much 

 more resistant than that obtained from the rabbit. 



The Microscopic (Orientation) Agglutination Test. 



This method is especially of use, when only small amounts of culture 

 or serum are obtainable. Also, if agglutination is employed for the quick 

 recognition of bacteria, as for example, when it is desirable to know whether 

 a blue colony on a Conradi-Drigalski-agar plate is typhoid or not. 



In such a case a drop of the immune serum in the dilution of i : 50 or 

 i : 100 is placed upon a cover-glass held with a Cornet's forceps, and a 

 small part of the bacterial colony for identification carefully mixed with 

 this serum. As controls, a mixture is made with salt solution and with 

 normal serum. If agglutination occurs, small granules or clumps can read- 

 ily be seen with the naked eye by holding up the cover-glass against the 

 light. The control glasses on the other hand should show only a homo- 

 geneous turbidity. These changes are still more evident if the mixture is 

 examined microscopically in the form of a hanging drop. (Described, 

 p. 108.) 



