460 Tetanus. 



hours later. If 0.033 cc. (300 times the fatal dose) of a culture, 

 0.0001 cc. of which killed mice inside of two days, is injected 

 into mice with 5 times the quantity of immune serum (0.2 cc.) 

 the animals will remain unaffected. The blood serum of rabbits 

 which were not immunized had no protective action. Similar 

 results were obtained by Tizzoni & Cattani in mice and rats 

 (1891) with blood of artificially immunized pigeons and dogs 

 (1-2 drops of dog serum rendered ineffective 0.5 cc. of toxic 

 filtrate of cultures). 



These experiments formed the basis of the serum immuniza^ 

 tion and serum therapy, for which the serum of horses is now 

 exclusively used, since Schiitz found that horses and sheep 

 may be also successfully immunized against tetanus, and pro- 

 duce a potent immune serum. 



Preparation of Immune Serum. According to Behring's method, a virulent 

 bouillon culture is obtained, 0.75 co. of which is fatal for rabbits in 3 to 4 days, 

 and of this bouillon 80 cc. is mixed with 0.25%; 60 cc. with 0.175%; 40 ce. with 

 0.125% trichloride of iodine, while 20 cc. is used without any mixing. Passing 

 from the most attenuated to the least attenuated cultures the horses to be immunized 

 are treated Once in eight days with subcutaneous injections of 10-20 cc. and finally 

 with the pure culture, 0.5 ee. being injected at first and thereafter each five days 

 the dose being doubled for each injection. 



According to the procedure of Eoux & Vaillard immunization is carried out 

 with filtrates of 4 to 5 weeks old cultures, of which 1/400 cc. kills a mouse. At 

 first a filtrate attenuated with Lugol's solution is injected subcutaneously in 

 increasing doses and later the pure toxin, first injected subcutaneously, then intra- 

 venously or intraperitoneally. 



Standardization of Immune Serum. According to Behring's procedure the 

 potency of the serum is determined in such a manner that the smallest quantity of 

 serum is ascertained by inoculations on mice, which renders a certain quantity of 

 tetanus toxin of a known strength, ineffective (toxin and s^rum are mixed and 

 injected subcutaneously). As a test toxin a dry tetanus normal toxin (TeTN) is 

 used, of which 1 g. kills 150,000,000 g. of mouse weight, that is 10,000,000 mice 

 of .15 g. body weight each. As normal serum - (TeAN.^) is designated that serum 

 of which 0.1 cc. renders ineffective 0.03 g. of normal toxin; serum, of which 0.01 

 cc. exerts such an action, possesses 10 times the strength (TeAN^") etc. One cc. 

 of normal serum contains one antitoxin unit (A.-E.). In Germany only a serum 

 can be marketed which possesses not less than at least ten times the strength 

 (TeAN""), of which 1 cc. contains at least 10 A.-E., or 0.01 cc. neutralizes 0.03, g. 

 of normal toxin. 



Serum prepared by the Pasteur Institute of Paris renders ineffective 1 cc. of a 

 solution diluted in proportion of 1:100,000, which is 100 times the fatal toxin 

 quantity (=z 0.001 g. dried toxin). 



Both sera when dried in the vacuum are also used in the powdered form, 

 10 g. of the German preparation containing 100 antitoxin units, while of . the 

 Parisian preparation 1 g. corresponds to 10 ce. of the liquid serum. There is also 

 an antitoxin prepared by Tizzoni and Catanni on the market, Which is obtained 

 from dog and horse serum. 



The blood serum of highly immunized horses when injected 

 subcutaneously in sufficient quantities, protects animals against 

 the pathogenic action of a simultaneous tetanus infection or 

 an infection occurring shortly afterwards. The immunity is 

 produced immediately, but lasts only a short time, about 3 to 

 4 weeks in horses, and then diminishes gradually, finally dis- 

 appearing entirely. A simultaneous injection of immune serum 

 and toxin causes local tetanic symptoms, which however last 

 only a few days whereupon the animals recover. Similar results 



