1911.] On the Factors Concerned in Agglutination, 431 



All four tubes were incubated for six hours at 37° G. and then allowed to 

 stand for 12 hours in the cold room. A distinct turbidity formed in tube 1. 

 The contents of the remaining three tubes remained perfectly clear. 



The middle-piece or globulin solution obtained from sheep serum was shown 

 to have the same properties as the middle-piece solution obtained from 

 guinea-pig serum. It is proposed to supplement these experiments by 

 examining the properties of the globulin solutions of a variety of animals. 



Discussion of Results. 

 An agglutinating serum contains two factors, both of which are necessary 

 to agglutination. The one is the specific antibody, the other a precipitable 

 substance, probably of the nature of a globulin. By the interaction of 

 antigen and antibody the molecules of the precipitable substance are 

 aggregated on the surface of the blood corpuscle or bacterium which is to be 

 agglutinated. 



The amount of specific antibody necessary to produce agglutination is 

 probably minute, and, by diluting an antiserum, a dilution can be obtained 

 which contains sufficient antibody but not sufficient of the precipitable 

 substance. By adding to such a dilution of the antiserum a solution of the 

 precipitable substance, derived from normal guinea-pig serum, agglutination 

 can be effected. The amount of precipitable substance necessary to produce 

 the agglutination of sheep corpuscles appears to be considerably larger than 

 the amount required to agglutinate typhoid bacilli. The precipitable 

 substance is thermostable, it is present in heated normal serum, and it can 

 be precipitated from normal serum with a fraction of the serum globulin. 

 It can also be precipitated from a solution in normal saline by a suitable 

 combination of an antigen with its antibody. This precipitate is small and 

 does not become visible until the experiment has been incubated for 

 several hours. 



It is probable that agglutination is effected during the earlier stages of 

 the aggregation of the molecules of the precipitable substance, that is to 

 say, before the process has advanced to the stage when a turbidity is 

 visible. The precipitable substance is probably identical with "con- 

 glutinin." There is, however, this difference between the results obtained 

 in these experiments and the conglutination effects of Bordet and his 

 collaborators. 



In Bordet's experiments, conglutination was obtained by the interaction 

 of four factors, namely, the red cells, the heated antiserum, heated ox serum 

 (conglutinin), and complement. In the experiments described in this paper, 

 agglutination was effected by the interaction of three factors — the red cells 



