THE AGGLUTINATION REACTION 211 



of the flocculation rates of 20 precipitating antisera. Horse antisera, and some 

 rabbit antisera, gave the two optimal ratios, sometimes very close together. Other 

 antisera, mainly rabbit, gave only a constant-antibody O.R. ; the constant-antigen 

 O.K. was indefinite or absent, the speed of flocculation being approximately constant 

 for all antibody-excess mixtures. Horse antibody is more soluble than rabbit 

 antibody, and the failure of horse-antibody compounds to precipitate in antibody 

 excess may be due to this greater solubility (see Brown 1935). 



The Agglutination Reaction. 



The original description of the agglutination reaction by Gruber and Durham 

 (1896) was concerned with the flocculation of bacteria ; but any foreign cells— yeasts 

 and other fungi, red blood corpuscles coming from another species, etc. — will 

 stimulate the formation of agglutinins when injected into the tissues, and will be 

 specifically flocculated in the test-tube under the influence of the antiserum so 

 produced. 



The reacting system differs from that concerned in precipitation, in that the 

 antigen is not in colloidal solution but forms part of the structure of an organized 

 cell, usually part of its surface. It is the behaviour of the sensitized cells that we 

 observe, not that of the isolated antigen-antibody compound. As a result, our 

 system frequently becomes highly complex. It is not often that the surface of a 

 bacterial cell is characterized by a single antigenic component. In many cases 

 several different antigens are concerned ; and the injection of the bacterial cells 

 into a rabbit, or other animal, will induce the formation of a corresponding number 

 of different antibodies. The problem of the antigenic structure of bacteria is, 

 however, so important that it will be more convenient to deal with it in another 

 chapter. For the moment we may confine our attention to the agglutination 

 reaction as such, merely noting that the possible intervention of a multiplicity of 

 antigens and antibodies is one of the factors that has to be allowed for when trying 

 to interpret any experimental findings. 



When observing agglutination we can, if we choose, watch the formation of 

 bacterial aggregates under a microscope. This method was once commonly em- 

 ployed in diagnostic agglutination tests and still has its uses. But in quantitative 

 titrations the macroscopic method, in which bacterial flocculation is observed with 

 the naked eye, or with the aid of a hand lens, is greatly to be preferred. 



The reactions that determine the agglutmation of bacteria are essentially similar to 

 those that determine the formation of a precipitate when a soluble antigen reacts with its 

 homologous antibody. It was, indeea, in connection with agglutination that the essential 

 role of electrolytes was first clearly demonstrated (Bordet 1899). If bacteria are allowed 

 to react with an agglutinating antiserum in a salt-free medium there is usually no aggre- 

 gation of the bacterial cells. That the antibody under these conditions unites with the 

 antigen may, however, be shown by demonstrating its absence from the supernatant fluid 

 after centrifugation of a mixture in which bacteria are present in excess, or by adding 

 an electrolyte to the resuspended cells, when agglutination occurs (see also Joos 1901, 

 1902, Bechhold 1904, Forges 1906, Forges and FrantschofF 1906). The concentration of 

 electrolytes is not without effect on the amount of antibody absorbed by bacteria or other 

 cells. In the analogous case of the binding of hsemolysin by red cells, or by red-cell stroma, 

 there is evidence that the amount of antibody bound at or about neutrahty (pH 7) is 

 greatly reduced in the absence of electrolytes (Coulter 1920-21, von Euler and Brunius 

 1931). Changes in pH also affect the absorption of antibody by antigen, particularly in the 

 absence of salts (Coulter 1920-21, von Euler and Brunius 1931, de Kruif and Northrop 

 1922-23). Many of these effects probably result from a change in the dissociation constant 



