THE TECHNIQUE OF SERUM REACTIONS 305 



lively little soagulable protein, and the quantity injected, even with 

 the largest tolerable doses of bacteria, may contain but very small 

 amounts of true protein forming antigen. 



The animals are weighed from time to time, and if considerable 

 loss of weight ensues, the intervals should be increased. Doses from 

 2 to 5 c.c. should be given. In giving the later injections the danger 

 of anaphylaxis must be remembered. A single injection of a large 

 quantity has occasionally yielded a precipitating scrum of consider- 

 able strength, 4 but this method is not usually successful. Injections 

 are made at intervals of from five to seven days. Seven to twelve 

 days after the last injection the animals may be bled from the ear, 

 and a preliminary test made to ascertain the precipitating value of 

 the serum. If this is insufficient, more injections may be made. 

 Bleeding should be done 7 to 12 days after the last injection. Such 

 sera may be preserved in the dark and at a low temperature. If 

 a preservative is added, Nuttall prefers chloroform to the phenols, 

 because of occasional turbidity produced by these. Fornet and 

 Miiller 5 and others have recommended rapid methods of precipitin 

 formation by injecting relatively large amounts of the antigen daily, 

 for three or four successive days, bleeding on the fifth or sixth day 

 thereafter. This method has been followed by a number of workers 

 subsequently, and is often successful, sera of considerable titers 

 being obtained. In our experience, however, this method has not 

 shown itself to be entirely advantageous, since it is rare that a very 

 potent serum is so produced, and also animals bleed within a week 

 after repeated large doses, may show in their serum not only anti- 

 bodies but also residues of antigen, the two substances not united 

 within the animal body, but gradually uniting and forming precipi- 

 tates after the serum has been obtained and stored. 



This phenomenon of the simultaneous presence of antigen and 

 antibody in the circulating blood has been variously explained, the 

 view formerly held being that of von Dungern, who believed that 

 every injected antigen contained partial antigens, a, b, c, etc., each 

 of which produced its partial antibody, A, B, C, which, by being 

 present in unequal proportions, gave rise to the simultaneous 

 presence of perhaps a and B, etc. Our own opinion is 6 that the 



*Michaelis, Deut. med. Woch., 1902. 



5 Fornet and Muller, Zeit. f. Hyg., 66, 1910. 



6 Zinsser and Young, Jour. Exper. Med., 17, 1913. 



