Diphtheria Antitoxin. 



231 



100 (1847) 



Agglutination phenomena with diphtheria antitoxin. 



By P. J. MOLONEY and L. O. HANNA. 



[From the Research Division, Connaught Antitoxin Laboratories, 

 University of Toronto, Toronto, Canada.] 



In search for an in vitro test for diphtheria antitoxin the 

 following observation was made: (a) When an emulsion of the 

 diphtheria bacillus, Park 8, is mixed with diphtheria antitoxin, 

 allowed to stand at 37 0 C. for 1 hour, centrifuged, washed with 

 saline and re-suspended, it is no longer agglutinated by diphtheria- 

 agglutinating serum, (b) The organisms sensitized in this way 

 are inagglutinable by acid agglutination. 



In this test for the inhibition of acid agglutination the cells are 

 suspended in a buffer solution which gives a maximum agglutina- 

 tion with unsensitized cells. The point of maximum agglutination 

 for acids varies somewhat depending on the culture and the buffer 

 mixture, but for phthalate mixtures (Clark and Lubs) diluted 

 1-1 with distilled water it is about P H 4.2 for a three or four-day 

 broth culture of the Park 8 strain. 



To determine the specificity of the test for diphtheria antitoxin, 

 experiments were carried out along four different lines. 



1 . The following sera were used in place of diphtheria antitoxin 

 and the test carried through: normal human serum, positive T.B. 

 human serum, normal guinea pig, concentrated tetanus antitoxin, 

 fresh antitoxic serum, normal horse, normal sheep, normal rabbit. 



In the above experiment there was inhibition of agglutination 

 when the cells had been sensitized with antitoxic serum and with 

 tetanus antitoxin ; and when the other sera had been used there 

 was agglutination. A guinea-pig test with the tetanus antitoxin 

 showed that it contained diphtheria antitoxin, and it was subse- 

 quently discovered that this tetanus antitoxin was from a horse 

 which had previously been used for the production of diphtheria 

 antitoxin. 



When diphtheria bacilli are sensitized with diphtheria agglutin- 

 ating serum instead of antitoxin there is inhibition of agglutination 



