PRACTICAL APPLICATION 243 



tetanus, however, the serums are usually given until a therapeutic effect 

 is noted, regardless of the number of units administered. If it were 

 possible to determine quickly and accurately the amount of toxin in a 

 given patient, then neutralization could be accomplished along the same 

 lines that make this possible in the test-tube. The indications are to 

 administer at once sufficient antitoxin to neutralize all the toxin, giving 

 subsequent doses large enough to overcome the toxin as it is produced 

 until the focus of infection is removed. 



Antitoxin should be kept in a cold place and protected from air and 

 light. When this is done, they usually do not deteriorate more than 30 

 per cent, of their original strength, and often much less, within a year. 

 All manufacturers place a larger number of units in the container than 

 the label calls for, in this way allowing for the gradual loss in strength up 

 to the date specified on the label. According to Park, the antitoxin in 

 old serum is just as effective as that in fresh serum, except that there is 

 less of it. 



PRACTICAL APPLICATION 



The employment of antitoxic serums both in prophylaxis and in the 

 treatment of infection, is considered in greater detail in the chapter on 

 Passive Immunization and Serum Therapy. 



Romer's Method of Determining Small Amounts of Diphtheria Antitoxin. The 

 principle of this method is based upon the observation that, when very small amounts 

 of diphtheria toxin are injected intracutaneously into the abdominal skin of guinea- 

 pigs, small areas of edema and necrosis result in about forty-eight hours. When 

 such injections are made with mixtures of toxin and antitoxin, the presence of free 

 toxin is indicated by such tissue changes. It is chiefly used in determining the anti- 

 toxin content of human serums after active immunization with the toxin-antitoxin 

 mixtures of von Behring. (See p. 718.) 



Technic. I conduct this test in the following manner: The "limes-necrosis" 

 (L n ) dose of a toxin is first determined, which is the amount of toxin which, together 

 with T^ of a unit of standard antitoxin, will still produce a minimal amount of ne- 

 crosis in forty-eight hours after intracutaneous injection into .guinea-pigs. A series of 

 dilutions of the L+ dose of a toxin is made, ranging from 1 : 5 to 1 : 100, and 0.2 

 c.c. of each mixed with 0.2 c.c. of antitoxin so diluted that each 0.1 c.c. contains 

 T7 Vtf of a unit. These mixtures are made in small test-tubes, the cotton stoppers 

 paraffined, and the tubes incubated for three hours and placed in the refrigerator for 

 twenty-one hours, after which 0.2 c.c. of each is injected into guinea-pigs (prepared 

 by pulling out the hairs); several injections may be made in each pig. 



When the L n dose of the toxin has been determined this amount is mixed in a 

 similar manner with varying amounts of the patient's serum being tested. The 

 amount of serum just neutralizing the toxin contains -nyVzr of a unit of antitoxin from 

 which the amount of antitoxin per cubic centimeter of serum may be computed. For 

 example I have found that 0.003 c.c. of serum of a person reacting negatively to the 

 Schick test (p. 719) neutralized this amount of toxin; therefore each cubic centimeter 

 of this person's serum contained 0.33 unit of antitoxin. 



