ROLE OF INFLAMMATION I53 



ink diluted previously 1:2, 1:4, 1:6, 1:20, 1:40, 1:80, and 1:100. The ink was 

 diluted with distilled water and filtered before using. Twenty-four hours 

 after the skin injections, each rabbit received a single intravenous injection 

 of 1 c.c. per kilo of body weight, of "A^^" previously diluted 1:20. The re- 

 actions were read four and twenty-four hoius after the intravenous injec- 

 tions. Not less than 3 rabbits survived in each group. They all showed severe 

 hemorrhagic necrosis in the areas prepared with "A^j^" but the B. typhosus 

 reacting factors had no \isible effect upon the skin sites injected with the 

 various non-bacterial substances. 



Another experiment was done in which rabbits were injected intrader- 

 mally with sodium arsenate, India ink and normal horse serum in various 

 dilutions as above. Four hours after the rabbits were injected intravenously 

 with "Aji" (1 c.c. of "Ajj^" diluted 1:20 per kilo of body weight). No skin 

 reactions followed the intravenous injection. 



In experiments conducted similarly to the foregoing, no skin-preparatory 

 factors were found in numerous batches of normal and immune sera of 

 various animals obtained during different stages of immunization, and sera 

 of rabbits injected with large doses of toxic filtrates some hours before bleed- 

 ing, all of which were collected under sterile precautions and properly pre- 

 served. There were also tested various batches of chicken plasma, i.e., normal 

 plasma; plasma of chicken immimized intravenously with Streptococcus viri- 

 dans cultine filtrate for fi\'e weeks and obtained t^venty-four hours after the 

 last injection; plasma of chicken immunized intiavenously with large doses 

 of B. typhosus "agar washings" filtrates for five weeks, obtained twenty-four 

 hours after the last injection. The results were also entirely negative. Skin- 

 preparatory and reacting factors were encountered in only one immune 

 horse serum which was stored in the laboratories for two years, and although 

 it was found sterile at the time of the test, it might have been contaminated 

 some time during its prolonged storage and resterilized by the preser\ative. 

 Chemically concentrated sera may also occasionally serve as skin-preparatory 

 factors. It is clear, however, that they are usually grossly contaminated during 

 the process of chemical fractionation and purification. As a matter of fact, 

 cultures made of a serum at various stages of chemical treatment consistently 

 showed B. proteus, B. coli, B. sub til is. staphylococcus, etc., prior to the addition 

 of the preservative. 



Skin-preparatory potency of mixtures of animal protein antigens 

 with antibodies: 



The antibodies employed were anti-horse rabbit sera, anti-human horse 

 sera, anti-human rabbit sera, and anti-horse goat sera. 



Each rabbit was prepared by a single intradermal injection of 0.25 c.c. of 

 the preparation tested and twenty-four hours later injected intravenously 

 Avith 50 reacting units of B. typhosus. The preparations were precipitates 

 and supernatant fluids deri\ed from mixtures of sera used as antigen and 

 sera containing the antibodies. The mixtures were made in proportions of 0.9 

 c.c. of serum containing the antibodies to each of the following amounts of 

 serum antigen: 0.025 c.c, 0.05 c.c, and 0.075 c.c of dilution 1:16; o.i c.c. of 

 dilution 1:2; and 0.10 c.c. of undiluted serum. The results which represent 



