216 THE ANTIGEN-ANTIBODY REACTIONS 



The Reversibility o£ the Union between Antigen and Antibody. 



The formation and precipitation of insoluble antigen-antibody complexes 

 imply a certain degree of irreversibility, a degree which is illustrated by the 

 Danysz phenomenon referred to above. Danysz (1902) showed that if a constant 

 amount of diphtheria toxin is added to a constant amount of antitoxin, the toxicity 

 of the mixture varies according to the way in which the addition is made. If, 

 for instance, to a given amount of antitoxin, the equivalent amount of toxin is 

 added all at once, the mixture is non-toxic. If the same amount of toxin is added 

 in two or more fractions, with an interval of 15 minutes or more between each 

 addition, the mixture will be highly toxic. The fraction of toxin added earlier 

 unites with more than its equivalent amount of antibody, so that there is insuf- 

 ficient antibody to neutralize the later fractions, and there is, during the period 

 of the experiment, no detectable redistribution of antibody among the toxin 

 molecules. The solubility of the compounds formed in antigen or antibody excess, 

 however, suggests that if the union was reversible, a precipitate formed from 

 equivalent amounts should dissolve in excess of antigen or antibody. The evidence 

 on this point is conflicting. 



Little solution in these circumstances could be detected in precipitates of the following 

 antigens with their respective antisera : Types I and II pneumococcal polysaccharides 

 (Sobotka and Friedlander 1928) ; haemoglobin (Breinl and Haurowitz 1930). On the 

 other hand, the following specific precipitates are soluble : diphtheria toxin in antibody 

 and antigen excess (Healey and Pinfield 1935) ; Type III polysaccharide (Heidelberger 

 and Kendall 1935a) ; hsemocyanin (Hooker and Boyd 1936), and ovalbumin (Boyd 1940). 



Boyd found that the ovalbumin precipitates were as readily soluble after 10 months' 

 storage in the cold. In other cases the antigen-antibody union appears to become less 

 reversible with time. For instance, both Enders and Shaffer (1937) and Morris (1940) 

 found that dilute mixtures of living Type I pneumococci and protective antibody were 

 at first infective, but became less so if the mixtures were allowed to stand before injection 

 into mice. It should be noted that the fact of decreased dissociability with age is not 

 necessarily due to increasing firmness of combination ; an irreversible denaturation of 

 the antibody protein in the compound may take place (see also Chapter 11, p. 342). 



The solubility of antigen-antibody complexes in high salt concentrations and, 

 in some cases, in antigen or antibody excess, suggests that dissociation of the 

 complex takes place. The establishment of this fact is important in deciding 

 which of the better-known chemical and physical processes best describes the 

 phenomena of precipitation and agglutination. With Bordet, we may interpret 

 these reactions as adsorption phenomena, or with Arrhenius and Madsen, as obey- 

 ing the mass law. In the equations both for the mass law and the adsorption 

 isotherm of Freundlich (see below) the amount of absorption complex, or of com- 

 pound, present in the system at equilibrium depends on the concentration of one 

 or more of the reactants. However, in most precipitin systems studied (see p. 219) 

 the composition and amount of the antibody-antigen complex is dependent, not 

 on the final, but on the initial concentration of the reactants, a fact that implies 

 a relatively high degree of irreversibility of the antigen-antibody union. The- 

 union takes place probably within a few seconds, in systems that precipitate 

 visibly in a few minutes (see Burnet 1931, Boyd and Hooker 1938, FoUensby and 

 Hooker 1939, Heidelberger, Treffers and Mayer 1940, and Boyd, Conn, Gregg, 

 Kistiakowsky and Roberts 1941). In this case the laws will apply only in the 

 earlier stages of the reaction and their ap])lication to data obtained mainly from 



