i 
1908.) Diphtheria Antitoxin. 405 
serum in four parts by the gradual addition of ammonium sulphate to half 
saturation. He found no accumulation of antitoxin in any one fraction of 
protein precipitated, the result indicating that the antitoxin was not carried 
down in a purely mechanical way. 
The limits for the precipitation of antitoxin by ammonium sulphate were 
determined. It was found that the initial precipitation of the antitoxin was 
practically coincident with the first precipitation of the proteins of serum, 
and was complete only after about 22 per cent. of ammonium sulphate had 
been added. If these points be examined on the ammonium sulphate pre- 
cipitation curves, it will be found that they are practically coincident with 
the alcohol precipitation limits. 
(C) By Neutral Salts in the Presence of Acids.—The precipitation of anti- 
toxin by salts in the presence of acids was examined; 5 per cent. acetic acid 
and 0:25 per cent. hydrochloric acid were found to have no deleterious 
influence on diphtheria antitoxin at room temperatures. Controls showed 
that these acids in dilutions corresponding to the antitoxic value of the 
serum used did not destroy toxin. The precipitation of antitoxin by sodium 
chloride and 0°25 per cent. hydrochloric acid started when about 10 per cent. 
of the total protein of the serum was precipitated, ~.e., the first 10 per cent. 
of protein precipitate contained only a minimal amount of antitoxin. 
During the rest of the protein precipitation the antitoxin came down in a 
quantity proportional to the amount of protein precipitate produced. There 
was no indication of any differentiation of the antitoxin from the great bulk 
of the proteid precipitate. 
(D) The Mechanical Precipitation of Diphtheria Antitoxin from Serum.— 
It has been stated above that the temperature at which antitoxin is destroyed 
differentiates it from ferments. | 
A few experiments were made to determine whether it possessed the 
property common to ferments of mechanical precipitation. Various flocculent 
substances precipitated in serum did not diminish its antitoxic properties. 
This absence of mechanical precipitation might have been due to the simple 
nature of the precipitants, e.g.,calcilum phosphate and kieseleuhr. <A solution 
of colloidal ferric phosphate was therefore added to antitoxic serum. The 
colloidal ferric phosphate was precipitated by the ionised salts present in the 
serum. After it had settled to the bottom of the flask as a gelatinous mass, 
the antitoxic value of the remaining serum was determined. There was no 
diminution in its antitoxic properties. From these experiments it may be 
concluded that diphtheria antitoxin does not possess the property of 
associating itself with substances precipitated in a solution of it. 
