484 



Journal of Agricultural Research 



Vol. XIII, No. lo 



An example; of a test. — Experiment 5. On May 4, 191 7, mixtures 

 containing tetanus serum 374 were prepared as detailed in Table III. 

 The same mixtures were used in experiments 4, 5, and 5.1. Chemical 

 data were obtained 6 and 17 days after digestion began. On the day of 

 the inoculation, 18 days after digestion began, a 0.5-c. c. portion of each 

 mixture was diluted to 20 c. c. with salt solution, resulting in five dilute 

 antitoxin solutions, all of which contained i unit per cubic centimeter 

 if none of the antitoxin had been destroyed. Of course, no antitetanus 

 units v/ere present at any time in mixture E. Measured quantities of 

 these diluted mixtures were then mixed with salt solution and tetanus- 

 toxin solution as detailed in Table IV, which is a typical one. As 

 three guinea pigs were used on each dose, 3 or 4 doses were prepared; 

 later but 2 guinea pigs on each dose were used, beginning with experi- 

 ment 6. 



Table IV. — Co^nposiiion of the doses injected {experiment 5) 



[o 50 c. c. of digestion mixture was diluted to 20.0 c. c. with physioloeical salt solution; i c. c. of diluted mixture 

 contained i unit of antitoxin if none had been destroyed] 



Mixture. 



Dose 



originally 

 contained. 



Unit. 



Mixture A 



Mixtures B, C, D 

 Mixture E 



07 

 10 

 20 

 10 

 20 

 40 

 80 



a In all fcxpeiTmcnts after this, but 0.02 c. c. of toxin solution was mixed in the dose of mixture E for i 

 guinea pig. 



It was possible that the trypsin and sodium carbonate or pepsin and 

 hydrochloric acid at their final dilutions might destroy 50 or 60 MLD 

 (minimal lethal dose) out of the 100 in the test dose, and still the mixture 

 E guinea pigs would die within the standard time of 96 hours. But by 

 giving this group of pigs but 2 MLD instead of 100, any destructive 

 action on the toxin would have manifested itself. Out of 12 such 

 guinea pigs in 6 experiments, 9 died within 96 hours, i within 120 hours, 

 I within 7 days, and i lived, indicating that there was no significant 

 destruction of toxin. 



Two samples of tetanus toxin, labeled "Fi " and "F4," were obtained 

 from the Hygienic Laboratory of the United States Public Health 

 Service. The test dose (100 MLD) was the same for both, 0.0007 grn. 

 The toxins v/ere kept in a Hempel sulphuric-acid desiccator evacuated 

 to 3 mm. of mercury, in a refrigerator room at 1° C, in the dark. 



The toxin and antitoxin were not mixed in separate syringes as prac- 

 ticed by Rosenau and Anderson; instead they were mixed in weighing 



