I074 VENOMS AND ANTIVENINS 



method is that it produces a very marked degree of immunity in the animal which 

 then yields highly antitoxic sera. Nevertheless, it has the disadvantage of taking too 

 much time (at least eight to twelve months). 



3. Neutral-mixture method. — In order to overcome this disadvantage, one can 

 resort to the method of neutral mixtures as used by Roux and Babes in France, 

 Kretz, Pawlowsky, and Maksutofif in Germany, Smith and McClintock and Ferry in 

 this country in the immunization against bacterial toxins. When applied to anti- 

 venins, this method requires only about three to five months to immunize a horse. 

 The first dose is two hundred to four hundred times as high as that used in the 

 mithridatization. This method, though not economical, because it consumes large 

 amounts of antitoxin, is practically harmless to the animals. 



4. Mixed method. — The immunization is begun with neutral mixtures, and the 

 dose of antitoxin is gradually decreased until this becomes so small that it can be 

 omitted and pure venom solution can be injected. By this method a horse can be 

 immunized in about four to six months and with considerable reduction of expense. 



ANIMALS USED 



Horses are the best animals for the immunization against snake venoms because 

 of their docility, resistance to venoms, and ready production of antibody in high con- 

 centration. Mules may also be used, but they are not so docile and resistant, and do 

 not produce antibodies in such high amounts nor so easily. Cows give antivenins of 

 low antitoxic value. Goats can be used in countries, such as some African districts, 

 where horses, mules, and cows are unavailable. They are quite resistant, but un- 

 fortunately they give only about 500 cc. of blood at each bleeding, while horses yield 

 at least 6-8 liters, mules 3-4 liters, and cows 4-6 liters. 



ANTIGEN PREPARATION 



Antigen preparation is one of the most important steps in the immunization 

 technique. In order to produce a highly active and efficient antivenin, the venom 

 must never be treated by any chemical or physical process which may affect its com- 

 position. It must retain all of its antigenic principles, and must be prepared in such 

 a way as to remain active indefinitely. 



The technique I use in order to accomplish this is as follows: Every fortnight the 

 venom of the specimens of each species is extracted from the glands and collected into 

 a wine glass, as shown in Figure i. After the extraction the venom is desiccated in 

 an incubator and afterward dissolved in equal parts of salt solution and glycerin 

 and used for inoculations. The solution must be about neutral, otherwise the venom 

 will gradually lose its activity. It must also be kept in the icebox about one month 

 in order to become sterile (ripening). 



VENOM PURIFICATION (STANDARD VENOM PREPARATION) 



It is known that venoms normally have a definite and invariable activity and, 

 therefore, can be used in the most accurate titrations. For this purpose, however, 

 they must be purified. This is done by centrifugalizing, decanting, and desiccating 

 the supernatant fluid which is pure venom. After drying, the pure venom is kept in a 

 vacuum over calcium chloride or other dehydrating substance. 



