1911.] On the Factors Concerned in Agglutination. 427 



precipitated by diluting a serum with distilled water .and saturating the 

 mixture with carbonic acid. In these characteristics it conforms to the 

 description of conglutinin which is given by Bordet and Streng. The substance 

 may be neither more nor less than serum globulin or some fraction of globulin 

 which is easily precipitated. 



The experiments suggest that an agglutinating serum contains a specific 

 antibody and a non-specific substance, both of which are necessary to produce 

 agglutination. When such a serum is greatly diluted the dilution may 

 contain sufficient of the specific antibody but not sufficient of the non-specific 

 anti-substance. In such a case the non-specific substance can be supplied by 

 adding middle-piece solution prepared from normal serum, and agglutination 

 is produced. 



Experiments to Determine the Way in which the Middle- Pieec Solution aids in 

 Agglutination. 



The inter-relation of the factors concerned in agglutination is to some 

 extent illustrated by the following experiment. Ten cubic centimetres of a 

 1 in 20 suspension of washed sheep red corpuscles were mixed with 20 c.c. 

 of a 1 in 200 dilution of a heated hemolytic antiserum. The mixture was 

 allowed to remain in the cold room for one hour and was then centrifugalised. 

 The corpuscles were then thoroughly washed and again centrifugalised. The 

 sensitised corpuscles were then mixed with 10 c.c. of middle-piece solution. 

 In another tube an equal quantity of unsensitised sheep corpuscles was added 

 to 10 c.c. of middle-piece solution. Both tubes were placed in the cold 

 room. At the end of this time very marked agglutination had taken place in 

 the tube containing the sensitised corpuscles. The contents of both tubes 

 were centrifugalised. The deposit of agglutinated cells was shaken up in 

 normal saline and again centrifugalised. After two washings the deposit 

 was thoroughly shaken up and suspended in normal saline. Within 

 15 minutes the cells had re-agglutinated, and after half an hour had fallen 

 in a compact mass to the bottom of the tube. 



The supernatant fluid from the tube which contained the sensitised red 

 cells was added to a further quantity of sensitised red cells. After the lapse 

 of several hours a very slight degree of agglutination could be detected. It 

 is evident that sensitised red cells can, in the process of agglutination, remove 

 from the solution of middle-piece the substance which produces the agglu- 

 tination. The supernatant fluid from the tube which had contained the 

 unsensitised red cells contained the agglutinative substance in a state of 

 unimpaired activity. Some substance had been taken out of solution by the 

 sensitised but not by the unsensitised e< lis. 



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