230 IMMUNOLOGY 



of toxins and antitoxins to be discussed in the next chapter, it 

 might be well to consider briefly the question of optimal propor- 

 tion of reagents and a few other factors. 



Proportion of Immune Serum and Antigen Used by Nuttall. 

 — Nuttall (1901, 1904) realized that certain ratios of immune 

 serum to antigen favor the formation of immune precipitate. In 

 his work he used ratios varying from 20 :1 to 200 :1 and higher. 

 Danysz (1902) observed the formation of a precipitate when 

 ricin was completely neutralized by antiricin. He found that the 

 ratio of ricin (toxin) and antiricin (antitoxin) that gave the 

 most voluminous precipitate also was the ratio necessary for 

 complete neutralization of toxin by the antitoxin. In 1909 Cal- 

 mette and JNIassol obtained similar results with venom and anti- 

 venom. They concluded that this offered a reliable method for 

 measuring tlie antitoxin content of immune serum. While a great 

 many have offered experimental evidence to show that the toxin- 

 antitoxin precipitation is due to bacterial protein and its bac- 

 terial precipitin, Bayne-Jones (1928) in an excellent review of 

 the literature as well as his own work concludes that the reaction 

 is between the toxin and antitoxin and not between bacterial 

 protein and bacterial precipitin. 



Importance of Optbial Proportions of Antigen and Antibody. 

 — In view of the importance which zoning plays in both the toxin- 

 antitoxin and precipitin reactions it would seem desirable to re- 

 view Dean's (1931) extensive investigations and also those of 

 others on the influence of optimal proportions of antigen and 

 antibody in the latter reaction. Dean found that when he mixed 

 e.g. a 1:5 dilution of mitihorse serum with varying dilutions of 

 antigen (horse serum), the largest amount of precipitate occurred 

 with a 1:8 dilution of antigen. When he used a 1:10 dilution 

 of antiserum, the largest precipitate occurred in the tube to 

 which he had added a 1 :16 dilution of antigen, etc. He obtained 

 similar results with antit>T)hoid serum and typhoid filtrates. He 

 also noted that when either antiserum or antigen was present 

 in relative excess the reaction was delayed. He says that 

 the importance of optimal proportions has been confirmed by Opie 

 (1923), Parker (1923) and Morgan (1923). The latter worked 

 with the soluble specific substance of the pneumococcus and the 

 homologous type of pneumococcus immune serum. More recently 



