REACTIVITY TO VARIOUS MICROORGANISMS 35 



saiy for elicitatioii ot the phenomenon with the above described 

 "aoar ^\'ashinos" filtrates. 



Cohn (1931) failed to obtain any reactions \vith filtrates of 

 10 per cent serimi broth cultures incubated for five to seven days. 

 In contrast, filtered washings of young blood agar cultures gave 

 strong reactions in 20 per cent of a small group of rabbits tested. 



In recent investigations Morimoto (1935) washed w'ith saline 

 twenty-four hour old cultures of gonococcus on blood agar. The 

 washings were heated at 60° c. for thirty minutes, frozen at 70° c. 

 for twenty-four hours and incubated at 37° c. for t^venty-four 

 hoins. The procedine of freezing and incubation was repeated 

 foiu' times. The results were positive. Hormone broth cultines 

 also yielded reactions. Older ctilttnes seemed to be more active 

 than young ones but no cpiantitative titrations ^vere reported. 

 This author filtered his material through Seitz discs. It should be 

 pointed out here that the discs have been sho^vn by Binnet 

 (1931) and others to reduce the potency of the principles. 



ACTIVE PRINCIPLES OF B. TYPHOSUS 



In the early experiments described in the introductory chapter, 

 active principles were deri\'ed from tryptic digest broth ctiltures 

 of a suitable strain of B. typlwsus. The skin-preparatory potency 

 of these materials titrated as high as 1:64 per 0.25 c.c. provided 

 at least 1 c.c. of the same undiluted material Avas employed for 

 the provocati\e injection. The reacting potency of filtrates of 

 tryptic digest broth cultines titrated against a single site prepared 

 with 0.25 c.c. of the imdiluted filtrate fluctuated between 50 to 

 200 units, per 1 c.c. (Method III) . 



Plain broth cidture filtrates yield poor preparations. In recent 

 work several batches of filtrates of three day old plain l^roth cid- 

 tures contained only 10 reacting iniits per 1 c.c. (Sh^vartzman, 



i935«) • 



Inasmuch as the abo\e materials, in addition to low potency, 

 also contained a large amount of extraneous substances derived 

 from the bacteria and the medium, the "agar ^vashings" filtrates 

 of B. typhosus were unquestionably the preparations of choice. 



B. typhosus "agar washings" filtrates: 



The "agar washings" filtrates are prepared essentially in the 



same manner as those of meningococcus (Shwartzman, 1929^) . 



Kolle flasks containing plain \cal infusion agar of pH 7.4 are 



