ANTIBODIES TO PNEUMOCOCCUS 375 



yielded specific precipitates, in dilutions of 1 to 1,000,000, with 

 Type III antipneumococcic horse serum, but failed to precipitate 

 homologous rabbit antiserum, giving rise to specific inhibition. 

 The authors found further that aldobionic acid, the structural 

 unit of Type III polysaccharide, did not precipitate homologous 

 antiserum. 



With Soo Hoo, Heidelberger and Kendall (1933) 628 described 

 a method, developed from the previous work of the two last-named 

 authors, for the microestimation of precipitin in antiserum. The 

 technique involved the use of a deep-red protein dye, the R salt of 

 azo-benzidene-azo-crystalline egg albumen. The procedure gave 

 the actual weight of precipitin and could be applied to the maxi- 

 mal amount of precipitable antibody in any antiserum. The ob 

 servation that four samples of immune rabbit serum produced in 

 response to injection of the dye contained over one hundred times 

 as much precipitin as the antigen injected, appeared to the au- 

 thors as supplementing the growing mass of evidence against the 

 theory that specific antigen fragments are actually incorporated 

 into the antibody molecule. 



In 1935, Heidelberger and Kendall 622 pointed out that the usual 

 immunological technique, namely, incubation of precipitin reac- 

 tions at 37° for two hours and allowing the tubes to stand in the 

 ice-box over night, while resulting in the maximal precipitation of 

 antibody from immune rabbit serum, in the case of antipneumo- 

 coccic horse serum or purified antibody, does not permit the estab- 

 lishment of a true equilibrium or the precipitation of the maximal 

 amount of antibody nitrogen. The authors recommended that 

 analyses of antipneumococcic horse serum should therefore be car- 

 ried out at 0° and the tubes containing the antigen and immune 

 serum should be allowed to stand in the cold for at least twenty- 

 four hours in order to ensure the completion of the reaction. 



In an accompanying paper, Heidelberger and Kendall 623 re- 

 ported the results of additional study of the quantitative relations 

 in the precipitin reaction between Type III polysaccharide and 



