262 PHARMACEUTICAL BACTERIOLOGY 



following is a brief outline of the manufacture of the marvelous remedy for 

 the treatment of the dread disease of childhood, namely diphtheria. 



i. Antidiphtheric Serum 



A. Selecting and Testing the Horse. Ordinary, normal, non-pedigree 

 horses are preferred, purchased under a guarantee of soundness. Even 

 though purchased under such a guarantee the animal is kept under ob- 

 servation for a few weeks and tested for glanders by the mallein test. 

 No animal is retained until it is proven that there is no latent or active 

 disease present. The animal is well housed and well cared for during the 

 entire time, under conditions as sanitary as it is possible to make them. 

 All laboratories are also regularly visited by a U. S. Government in- 

 spector, who reports his findings to Washington. 



B. Preparing the Toxin of Diphtheria. Pure cultures of a selected 

 strain of the diphtheria bacillus, possessed of a high potency, virulency or 

 toxicity, are made in liter flasks containing beef bouillon. The original 

 bacilli thus used are taken from some patient suffering with diphtheria, and 

 by means of isolation methods all foreign microbes are rejected or excluded. 

 After the culture is several days old or when a maximum amount of the 

 toxin has been formed and deposited in the bouillon, the bacili are killed by 

 adding 0.25 per cent, of trikresol. The bouillon with the dead bacilli is 

 filtered. The clear filtered substance constitutes the toxin which is in- 

 jected into the horse for the purpose of developing (in the horse) the anti- 

 toxin of diphtheria. The virulency or potency of the toxin varies and is 

 tested on guinea-pigs and compared with the U. S. Government standard. 

 The highiy toxic race or strain of germs is perpetuated in the laboratory 

 by daily transfers to new culture tubes. In this manner the bacilli are 

 maintained for a long time, several years or longer. However, even 

 with the greatest care the race finally deteriorates, weakens or undergoes a 

 change in potency and it becomes necessary to secure a new stock culture. 



C. Developing the Antitoxin of Diphtheria in the Horse. Twice weekly 

 the horse is given (by hypodermic injection into the flank region) gradually 

 increasing doses of the toxin of diphtheria. The rule is to give enough to 

 produce a marked reaction. For a day or two the horse is sick with diph- 

 theria, then recovers as the increased antitoxin in the blood (serum) of the 

 animal neutralizes the toxin. This is continued for from four to six weeks 

 when a maximum amount of antitoxin has presumably developed. The 

 last dose of toxin is several hundred times greater than the first. 



D. Bleeding the Horse. A sterilized canula or trochar is inserted into 

 the jugular vein, after the neck has been thoroughly washed with soap and 

 water, shaved and rinsed with a 5 per cent, solution of carbolic acid. The 

 blood is drawn off into sterilized liter tubes, which are plugged with cotton. 



