152 STUDIES IN IMMUNITY. 



tube is added distilled water and to the second normal salt solution. 

 After agitation these tubes are again centrifugalized. It is found that 

 the bacteria in the tube containing salt solution go to the bottom 

 much more rapidly than in the one containing distilled water. 

 When they are finally deposited the supernatant fluid is removed 

 from each tube and replaced by a second amount of the same fluid, 

 that is to say, salt solution in one and distilled water in the other. 

 The bacteria are then shaken up in the fluid. It is found that 

 clumps form rapidly in the tube containing salt solution, but that the 

 bacteria remain indefinitely in suspension in the tube containing dis- 

 tilled water. If a small amount of the cloudy fluid, say 10 c.c., is 

 taken from the second tube and placed in a fresh tube and to it 

 is added 0.07 grams of NaCl, agglutination reappears and the deposi- 

 tion of bacteria takes place.* 



The same phenomenon occurs with the cholera vibrio and a 

 normal agglutinating serum in place of a specific cholera serum. 

 We noted three years ago that normal horse serum agglutinates 

 the cholera vibrio and other bacteria such as B. typhosus, B. coli 

 and B. tetani very markedly. If this last experiment is repeated 

 with normal horse serum instead of the specific serum the same 

 results are obtained. 



In this latter case it is not necessary to remove all the traces of 

 NaCl by repeated washing. The salt solution containing the 

 clumped bacteria is simply centrifugalized and the supernatant 

 fluid decanted; the deposition is then divided into two parts and 

 placed in two separate tubes ; one tube is filled with distilled water 

 and the other with salt solution. On agitation the agglutination 

 recurs only in the presence of salt.f A similar experiment with 

 normal horse serum and B. typhosus gave the same result. 



* It must be noted that this."reagglutination" of bacteria on the addition of 

 salt to distilled water does not occur quite so rapidly as in the tube in which the 

 bacteria have remained in contact with salt solution, especially when the contact 

 with distilled water has been prolonged. It is probable that the micro-organisms 

 must retain a certain amount of salt in order to agglutinate well. 



t The presence of an agglutinating power in serum has doubtless vitiated many 

 researches on the bacterial power of body fluids. Many observers, indeed, have 

 used the method of inoculating at intervals small amounts of a mixture of serum 

 and bacteria in gelatin in order to determine the destructive power of the serum. 

 It is quite possible that the serum in question, when the amount of bacteria is small, 

 may so clump them that each clump of bacteria will give rise to a single colony 



