70 TOXIN AND ANTITOXIN. 



Guinea-pigs surviving a dose of toxin, may after two to four weeks begin to show 

 paresis first of the hind, then of the fore extremities, and finally even of the muscles of 

 the back and respiration. The most severe types of such conditions, however, may fully 

 subside. They may be considered as analogous to the post-diphtheritic paralysis taking 

 place in man, which is usually, as is well known, of a benign nature. 



Besides guinea-pigs other animals suitable for diphtheria experimental work 

 are rabbits (especially by intravenous injection) and pigeons (by intramuscular 

 injection). 



The susceptibility of animals towards diphtheria toxin varies greatly, as is seen from 

 the following scale of Behring, the least susceptible animals being mentioned first: 

 mouse, rat, dog, guinea-pig, rabbit, sheep, cow, horse, goat. 



The strength of the diphtheria toxin is estimated as follows: 



Estimation of Guinea-pigs of equal size (250 gms.) receive subcutaneous 



Strength of injections of decreasing amounts of toxin. With a strong 



Diphtheria toxin, centi- and milligrams or even smaller denominations 



Toxin. are O f sufficient potency to produce death. Doses such as 



these are not injected unless diluted in normal salt solution. 



For exact results one must not depend upon the findings brought out by 



the injection of a single animal with each dilution; several should be 



inoculated with the same dose and the effects, which should be the same in 



all cases, noted. It is impossible to state beforehand how many dilutions 



may be necessary. If the various actions, dependent upon the successive 



gradations of dosage are successfully represented, the experiment may be 



taken as conclusive; that is to say, the smallest doses must leave the animal 



entirely unaffected, the moderate produce slight local and general symptoms, 



and the larger ones cause death of the animals. If it should so happen 



that they all die, a new set of experiments employing a lower scale of dosage 



should be undertaken. 



Thus it is seen that the action of diphtheria toxin is subject to the quantity 

 of the toxin injected. If several different diphtheria toxins are tested at the 

 same time, it is at once evident what far reaching differences may arise. 

 While o.ooi c.c. of one diphtheria toxin kills a guinea-pig in twenty-four 

 hours, a different diphtheria toxin performs the same action with a dose ten 

 times as great, e.g., o.oi c.c. The second toxin thus contains only one- 

 tenth as many of the active substances. In order to obtain a uniform method 

 for estimating the strength of a diphtheria toxin and thus get comparative 

 values, a standard unit of the same has been adopted. And this consists of the 

 smallest amount of toxin that will kill a healthy guinea-pig weighing about 250 

 gms. in four to five days. This is known as the minimum lethal dose or dosis 

 letalis minima. In addition to this "direct toxic value," it is frequently 

 important, especially for the standardization of curative sera, to estimate 

 the "indirect toxic value" by which is meant the amount of antitoxin which 

 a toxin can bind or neutralize. 



