286 AGRICULTURAL AND INDUSTRIAL BACTERIOLOGY 



mine its antitoxin content. The horse is bled from the 

 right jugular vein by means of a sterile trocar and tube into 

 a sterile glass vessel. Usually about one liter of blood may 

 be secured per one hundred pounds weight of horse. This 

 amount can be withdrawn from the animal without serious 

 injury. After a period for recuperation the animal may 

 then be injected several times more with diphtheria toxin 

 and again bled. Successive bleedings from the same animal 

 may thus be secured over a period sometimes of several 

 years. The blood is allowed to clot, that is, the fibrinogen 

 of the plasma is transformed into fibrin which catches the 

 blood cells as in a net and by its contraction squeezes out the 

 blood serum from the clot. This clear, straw-colored blood 

 serum is found upon examination to contain the antitoxin. 

 Standardization of Antitoxin Serum. Inasmuch as 

 there is great variation in the relative antitoxic content of 

 the serum secured from different horses it is necessary to 

 standardize each sample. For this purpose, the antitoxin 

 content is compared with a sample of standard antitoxin, 

 secured in the United States from the Hygienic Laboratory 

 of the Public Health Service in Washington, D. C. The 

 standard serum secured from this source is diluted so that 

 each cubic centimeter contains one immunity or antitoxic 

 unit. It is evident that the antitoxin which has been 

 manufactured cannot be compared directly with the stand- 

 ard antitoxin but only by use of titration of each of these 

 against a toxin. The manufacturer, therefore, uses a suit- 

 able sample of toxic broth. This is first titrated against the 

 standard antitoxin secured. To each one of a series 

 of tubes one antitoxic unit of the standard serum is 

 added. To each of these tubes, then, graduated amounts 

 of the toxic broth are likewise added; to the first 

 tube a small amount, to the next a larger amount and 

 so on to the last. Each tube is then made up to the 

 same volume with sterile physiological salt solution and the 



