APPENDIX 463 



All operations in bleeding" the animals and preparing the 

 sera should be performed with precautions to avoid microbial 

 contamination. If the sera become contaminated they may be 

 sterilized by centrifugation followed by filtration through 

 ultrafine sintered glass or Berkefeld filters. It is most unwise 

 to add preservatives because this will interfere with certain uses 

 of the sera. For instance, the presence of 1 part in 10,000 of 

 Merthiolate in a serum which was then diluted 1 :100 for use in 

 one-step growth experiments caused a striking lengthening of 

 the latent period of phage multiplication. 



Assay of Antiphage Sera 



The reaction between the phage and its antibody is not 

 instantaneous, instead proceeding at a readily measurable 

 rate. This rate is a function of the concentration of virus, of 

 the concentration and activity of the antibody and of the tem- 

 perature. At constant temperature the rate may be expressed 

 as 



~dp/dt = Kp/D (differential) 



K = 23 D/t X log po/p (integral) 

 p/pQ = e"^^'^ (exponential) 



in which pQ = phage assay at zero time, p = phage assay at 

 time t min., D = final dilution of serum in the phage-serum 

 mixture; i.e., if the concentration is Vioo that of the undiluted 

 serum, then D = 100, K = velocity constant. 



This equation holds for inactivation of 90-99 per cent of the 

 phage present in the reaction mixture. The value of /T is a 

 characteristic of the particular lot of serum used. Once the 

 A' value of a sample of serum has been determined, it can be 

 substituted in the integral form of the equation and used to 

 calculate the dilution D of that serum to be used to inactivate 

 any desired fraction of the phage, e.g., 90 per cent, in any given 



