238 ANTITOXINS 



soon as the temperature reaction has subsided, a second subcutaneous 

 injection of a slightly larger dose is given, the amount of toxin increasing 

 about 10 to 15 c.c. per dose until, six weeks later, the animal is receiving 

 from 20 to 30 times, and on the sixtieth day about 60 times, the amount 

 originally given. At the- end of this time a trial bleeding is made and 

 the serum tested. 



There is absolutely no way of judging which horses will produce the 

 highest grades of antitoxin. Roughly estimated, those horses that are 

 extremely sensitive and those that react feebly are the poorest, but 

 there are exceptions even in these cases. The only reliable method, 

 therefore, is to bleed the horses at the end of six weeks or two months 

 and test their serum. As shown by Park and Zingher, persons yielding 

 negative Schick tests respond to toxin-antitoxin injections with the 

 production of antitoxin more readily than persons who react in a positive 

 manner. Taking advantage of this data, Hitchens and Tingley have 

 injected 0.2 c.c. diphtheria toxin equal to 3 M.L.D. for 250-gram pigs 

 into the conjunctiva of one eye of horses under examination for the 

 purposes of immunization, and found that many yielded negative tests 

 read at the end of forty-eight hours, indicating the presence of natural 

 diphtheria antitoxin in the blood of normal horses; these animals are 

 probably to be preferred in the production of antitoxin, as they are 

 likely to yield highly potent sera. If only high-grade serum is wanted, 

 all horses that give less than 150 units per cubic centimeter should be 

 discarded. The remaining horses should receive steadily increasing 

 doses, the rapidity of the increase and the interval of time between the 

 doses (three days to one week) depending somewhat on the reaction 

 following the injection, an elevation of temperature of more than 3 F. 

 being undesirable. 



For example, according to Park, a horse that yielded an unusually 

 high grade of serum was started on 12 c.c. of toxin (xuir c.c. fatal dose), 

 together with 10,000 units of antitoxin. Sixty days later a dose of 675 

 c.c. was given, and the serum contained 1000 units of antitoxin per 

 cubic centimeter. Regular bleedings were made weekly for the next 

 four months, at the end of which time the serum had fallen to 500 units 

 in spite of weekly gradually increasing doses of toxin. At the end of 

 three months the antitoxic serum of all the horses should contain over 

 300 units, and in about 10 per cent, as much as 800 units in each cubic 

 centimeter. Not more than 1 per cent, give above 1000 units, and, 

 according to Park, so far none has given him as much as 2000 units per 

 cubic centimeter. The very best horses, if pushed to their limit, con- 



