482 BACTERIOLOGY. 



in 4 c.c. The test-dose of toxin is added to this amount of serum and 

 the mixture is then injected subcutaneously into a 250 g. guinea-pig. 

 If the trial serum actually does contain 1 I. U. death will not result 

 till, as in the above determination of the test-dose, on about the 4th 

 day. If the animal dies on the 2nd or 3rd day it is evident that the 

 amount of serum taken contains less than 1 1. U. and that the original 

 serum is of less strength than advertised. If, on the other hand, the 

 animal does not die till about the 6th day, or even recovers, it follows 

 that the amount of- serum taken contains more than 1 I. U. 



In the latter case a series of guinea-pigs can be injected with 

 mixtures of the test-dose of toxin and variable amounts of the dilute 

 serum. Eventually a dilution of the serum will be reached which will 

 correspond, as indicated above, to 1 I. U. The number of I. U. in 1 c.c. 

 of the undiluted serum can then be readily calculated. 



Immunization against Diphtheria. 



The diphtheria toxin prepared according to the direc- 

 tions given can be employed for immunizing several rabbits. 

 The animals should first be weighed and their temperature 

 taken. For the first injection they should not receive more 

 than what corresponds to the minimum fatal dose for guinea- 

 pigs. Indeed, this amount of toxin may be injurious and it 

 is advisable to add to the toxin a drop of diluted iodine solu- 

 tion (p. 288). The animals can tolerate the toxin, thus 

 modified, better than if it were used direct. The tempera- 

 ture and weight of the animals are taken on each following 

 day. When the condition of the animals has become nor- 

 mal, a second injection is made and this is repeated until 

 the animals cease to appreciably react. The dose of toxin 

 is then doubled and this amount with a little iodine is 

 repeatedly injected. A new injection should not be given 

 until the animals have fully recovered from the preceding one. 

 When the animals shows no marked results, variation in 

 temperature or loss in weight, after the injection of this 

 amount of toxin then 4 times the minimum fatal dose may 

 be used, mixed as before with a little iodine. Eventually 



