248 IMMUNOLOGY 



of toxins and antitoxins. He devised methods for the measure- 

 ment of each and formulated a theory to explain the phenomenon 

 of toxin neutralization by antitoxin. Ehrlich's original method 

 of determining the units of toxin and antitoxin are discussed briefly 

 in Chapter XI. 



In 1922 Bamon introduced a flocculation metJiod for the titration 

 of toxins and antitoxins. Since then, many papers have appeared 

 in which both the mechanism of the reaction and the significance 

 of the results are discussed. Two of the most important publica- 

 tions bearing upon the subject are those of Bayne- Jones (1924, 

 1928). He says that Danysz (1902) observed the formation of a 

 flocculent precipitate in neutral mixtures of ricin and antiricin. 

 Similar results were obtained by Calmette and Massol (1909) 

 with mixtures of venoms and antivenoms. Nicolle, Cesari and 

 Debains (1920) stratified antitoxin over toxin-gelatin mixtures 

 and noted the formation of a precipitate in certain tubes. Ramon 

 (1922) added a uniform amount of toxin and diminishing amounts 

 of antitoxin to a series of tubes and noted that a flocculent pre- 

 cipitate formed first in the tube containing equivalent propor- 

 tions of toxin and antitoxin. This tube he called the ''indicator 

 tube." If the strength of the antitoxin, i.e., units per cubic 

 centimeter, is known, one can readily calculate the amount of 

 toxin per unit of antitoxin in the indicator tube. The amount 

 of toxin which brings about the most rapid formation of a floc- 

 culent preeipitate when mixed with one unit of antitoxin is called 

 by Ramon the Lf or flocculating dose of toxin. A flocculent pre- 

 cipitate occurs later in other tubes containing either an excess or 

 an insufficient amount of antitoxin but the tube in which the 

 flocculent precipitate first forms contains a balanced mixture of 

 toxin and antitoxin. In the following protocol illustrating the 

 Ramon flocculation technique it will be observed that 3.0 c.c. of 

 toxin are added to each of 6 small test tubes. Antitoxin having 

 a strength of 300 units per cubic centimeter was added in amounts 

 varying from 0.18 to 0.06 c.c. The contents of the tubes were 

 mixed and the tubes were placed in a water-bath having a tempera- 

 ture of 54° C. In this particular experiment they were observed 

 at five-minute intervals for one hour. The results are tabulated 

 in Table VII. 



