98 BACTERIOPHAGES 



arate from phage lysates soluble substances possessing the 

 antigenic specificity of the phage particles. When antiphage 

 antibody is allowed to react with such substances it loses its 

 ability to neutralize phage. Some, if not all, phage-antiphage 

 systems fix complement. These and other properties of anti- 

 phage sera will now be discussed in more detail. 



1. Preparation of Antiphage Sera 



Rabbits are the most convenient animals to use for antibody 

 production, and most of the serological work with phage has 

 been done with rabbit sera. The phages themselves are non- 

 toxic and nonpathogenic for animals and can usually be in- 

 jected in large amounts without damage to the recipient. 

 Crude phage lysates, however, may contain toxic bacterial sub- 

 stances, such as the endotoxins of the enteric bacteria and the 

 much more potent exotoxins of the corynebacteria and Clos- 

 tridia. In such cases the toxins must first be removed by frac- 

 tionation or inactivated. For many purposes it is desirable to 

 purify the phages before using them for immunization, since 

 adequate purification will result in the absence of host-cell 

 antibodies from the resultant antisera (Kalmanson, Hershey, 

 and Bronfenbrenner, 1942). If crude phage preparations 

 are used it may be necessary to remove host-cell antibodies by 

 absorption with bacterial suspensions. This is essential, for 

 instance, in studies involving complement fixation. 



Stimulation of antibody production requires the administra- 

 tion of adequate quantities of antigenic material. The daily 

 injection of about 10^'' virus particles (about 10 jjig.) for five days 

 will usually result in the presence of considerable neutralizing 

 antibody in serum obtained one week after the last injection. 

 Repetition of this course of immunization will further increase 

 the antibody titer. The final result does not seem to depend 

 greatly on the route (intravenous, intraperitoneal, or subcu- 

 taneous) by which antigen in administered. Use of the sub- 

 cutaneous route, however, is least likely to lead to toxic or ana- 

 phylactic reactions. Since there are large differences in anti- 



