744 MICROBIOLOGY OF DISEASES OF MAN AND DOMESTIC ANIMALS 



date of from eighteen months to two years is applied to diphtheria 

 antitoxin. 



It has been demonstrated that the antitoxic content of serum is 

 closely associated with the globulins. Advantage is taken of this 

 fact by most laboratories in reducing the volume of antitoxin, or 

 concentrating the product, by precipitating the globulins with 

 ammonium sulphate, redissolving the precipitate and dialyzing. 

 The concentration of serum by this method increases the unit value 

 per volume and tends to decrease the occurrence of undesirable second- 

 ary effects ("serum sickness"). 



TETANUS ANTITOXIN. The processes involved in the preparation 

 of antitetanic serum differ but little from those employed in the 

 manufacture of diphtheria antitoxin. The pure culture of B. tetani 

 is inoculated into large flasks of glucose bouillon, placed under an- 

 aerobic conditions and incubated at body temperature. A convenient 

 method of excluding free oxygen, in the presence of which the tetanus 

 organisms will not multiply, consists in boiling the glucose bouillon 

 before the inoculation, to drive off the oxygen, then covering the 

 liquid medium by a layer of oil. These cultures are subjected to a 

 temperature of 37 for about two weeks, after which they are examined 

 microscopically, preservative is added and the organisms are removed 

 by nitration. On account of the presence of spores and the danger at- 

 tending the contamination of any materials or biological products with 

 the tetanus bacillus, it is important that great care should be exercised 

 in the filtration and preparation of the tetanus toxin. Therefore, the 

 nitration process is best accomplished in an isolated room which is 

 used only for the preparation of tetanus toxin. 



Tetanus antitoxin is produced by the injection of horses with the 

 specific toxin and the same general methods and precautions are ob- 

 served as in the preparation of diphtheria antitoxin. The anti- 

 tetanic serum is tested relative to potency, safety and freedom from 

 microbial contamination. *The standard unit of tetanus antitoxin is 

 regarded as ten times the least quantity of antitetanic serum necessary 

 to save the life of a 35o-g. guinea-pig for ninety-six hours, against the 

 official dose of a standard toxin furnished by the Hygienic Laboratory 

 of the Public Health Service. 



Tetanus antitoxin is put up for use in the same manner as diph- 



* See U. S. Treasury Department, Public Health Reports, Vol. XXIV, No. 20, 1904. 



