THE TOXINS OF DIPHTHERIA 409 



After injection either of the toxin or of the living bacilli, 

 when the animals survive long enough, paralytic phenomena 

 occasionally occur. The hind-limbs are usually affected first, the 

 paralysis afterwards extending to other parts, though sometimes 

 the fore-limbs and neck first show the condition. Sometimes 

 symptoms of paralysis do not appear till two or three weeks 

 after inoculation. After paralysis has appeared, a fatal result 

 usually follows in the smaller animals, but in dogs recovery may 

 take place. There is evidence that these paralytic phenomena are 

 produced by toxone, as they specially occur when there is injected 

 along with the toxin sufficient antitoxin to neutralise the more 

 rapidly acting toxin proper. This toxone is supposed by Ehrlich 

 to have a different toxic action, i.e., a different toxophorous 

 group (p. 195), from that of the ordinary toxin; it produces 

 the late nervous phenomena, while its local action on the tissues 

 is very slight. It also has a weaker affinity for antitoxin, and 

 thus much of it may be left unneutralised. It is to be noted in 

 this connection that paralytic symptoms are of not uncommon 

 occurrence in the human subject after treatment with antitoxin, 

 the explanation of which occurrence is probably the same as 

 that just given. One point of much interest is the high degree 

 of resistance to the toxin possessed by mice and rats. Roux and 

 Yersin, for example, found that 2 c.c. of toxin, which was 

 sufficient to kill a rabbit in sixty hours, had no effect on a 

 mouse, whilst of this toxin even y 1 ^ c.c. produced extensive 

 neltosis of the skin of the guinea-pig. 



Preparation of the Toxin. — The obtaining of a very active toxin in 

 large quantities is an essential in the preparation of anti-diphtheritic 

 serum. Certain conditions favour the development of a high degree 

 of toxicity, namely, a free supply of oxygen, the presence of a large pro- 

 portion of peptone or albumin in the medium, and the absence of substances 

 which produce an acid reaction. In the earlier work a current of sterile 

 air was made to pass over the surface of the medium, as it was found 

 that by this means the period of acid reaction was shortened and the 

 toxin formation favoured. This expedient is now considered un- 

 necessary if an alkaline medium free from glucose is used, as in this 

 no acid reaction is developed ; it is then sufficient to grow the cultures 

 in shallow flasks. The absence of glucose may be attained by the 

 method described above (p. 79), or by using for the preparation of the 

 meat extract flesh which is just commencing to putrefy (Spronck). 

 L. Martin uses a medium composed of equal parts of freshly prepared 

 peptone (by digesting pigs' stomachs with HC1 at 35° C), and glucose- 

 free veal> bouillon. By this medium he has obtained a toxin of which 

 sfar c.c. is the fatal dose to a guinea-pig of 500 grms. Park and 

 Williams and also Dean found that the amount of glucose present in 

 ordinary beef is not sufficient to interfere with toxin formation, provided 

 that a considerable amount of peptone, 2 per cent., be added, and the 



