352 PROTECTIVE INOCULATIONS. 
the rods, but from pus in which no spores are seen cultures of the 
bacillus may be obtained in which spores will develop in the usual 
manner. 
Guinea-pigs are even more susceptible to the tetanus poison than 
mice, and rabbits less so. The amount of filtrate from a slightly 
alkaline bouillon culture required to kill a mouse is extremely minute 
—0.00001 cubic centimetre (Kitasato). The tetanic symptoms are 
developed within three days; if the animal is not affected within four 
days it escapes entirely. The tetanus poison is destroyed by a tem- 
perature of 65° C. maintained for five minutes, or 60° for twenty 
minutes, or 55° for an hour and a half; in the incubating oven at 37° 
C. it gradually loses its toxic potency’; in diffuse daylight, also, its 
toxic power is gradually lost; in a cool, dark place it retains its origi- 
nal potency indefinitely ; in direct sunlight it is completely destroyed 
in from fifteen to eighteen hours; it is not injured by being largely 
diluted with distilled water; it is destroyed in an hour by hydro- 
chloric acid in the proportion of 0.55 per cent; terchloride of iodine 
destroys it in the proportion of 0.5 per cent; cresol in one per cent— 
one hour’s exposure. In general it is destroyed by acids and by 
alkalies. Blood serum from cattle, horses, sheep, rabbits, rats, or 
guinea-pigs does not modify its toxic properties. 
Brieger (1886) first succeeded in obtaining from impure cultures 
of the tetanus bacillus a crystallizable toxic substance, called by him 
tetanin, which was found to kill small animals in very minute doses 
and with the characteristic symptoms of tetanus. More recently Kit- 
asato and Weyl have obtained the same substance, by following 
Brieger’s method, from a pure culture of this bacillus. From a bouil- 
lon made from one and one-fourth kilogrammes of lean beef, with the 
addition of twenty-five grammes of peptone, they obtained 1.7118 
grammes of hydrochlorate of tetanin. This proved fatal to white mice 
in six hours in the dose of 0.05 gramme, and a dose of 0.105 gramme 
caused characteristic tetanic convulsions and death within an hour. 
The bacteriologists last named also obtained from their cultures the 
tetanotoxin of Brieger. Two mice were inoculated subcutaneously 
with 0.003 gramme of this substance; one died at the end of five 
hours without the development of tetanic symptoms; the other sur- 
vived. In addition to these substances, indol, phenol, and buty- 
ric acid were demonstrated to be present in cultures of the tetanus 
bacillus. 
The more recent researches of Brieger and Frankel, and of Kita- 
gato, show that the toxic ptomain discovered by Brieger in 1886 is 
not the substance to which cultures of the tetanus bacillus owe their 
