32 IMMUNO-CATALYSIS 



immunized with sterile filtrates obtained from B. enteritidis cultures 

 incubated in a synthetic medium for six to 14 days contained ag- 

 glutinins, precipitins and complement fixing antibodies. They could 

 not demonstrate the presence of protein in these filtrates by the biuret, 

 ninhydrin and Molisch tests; Millon's test for tyrosine and Ehrlich's 

 and vanillin tests for tryptophane were negative. 



Ten liters of toxic filtrate were concentrated in vacuo at 40°C. to a 

 syrupy fluid of 200 ml. The protein tests still were negative, although 

 the residue was still toxic for rabbits. After dialysis of the residue against 

 distilled water the dialysate was concentrated in vacuo at 30° to 40°C. 

 to a volume of about 5 ml. This concentrate showed by ninhydrin, 

 vanillin, and diazo tests (the last for histidine) very faint, but definitely 

 positive reactions. By comparing the sensitiveness of these tests to the 

 positive reactions shown by the concentrate they calculated the protein 

 content of the original solution at least as 0.000,000,1 g. per ml. of 

 filtrate. Rabbits on immunization with 30 to 40 ml. filtrates contain- 

 ing a total of 0.000,003 to 0.000,004 g. of protein elicited definite 

 antibody formation. The authors in the light of their work suggest that 

 such infinitesimal amounts should be borne in mind in interpreting 

 results obtained with apparently protein-free materials. 



Topley (1930) evaluated as nearly as possible the relation between 

 the amount of antigen injected into rabbits and the resulting agglu- 

 tinin titre. The antigenic material consisted of a saline suspension of 

 Bact. 'paratyphosum B. in the type phase, and the antibody studied 

 was the corresponding H or flagellar agglutinin. Bacterial suspensions 

 containing 0.25 per cent formalin were killed by heating at 55°C. for 

 one hour. A series of rabbits were injected intravenously with 10^, 

 10^ and 10^ bacilli per kilo body weight (k.b.w.); specimens of sera 

 were collected at intervals of three to seven days during the first few 

 weeks and at longer periods thereafter. The whole period of observa- 

 tions varied from 50 to 300 days or more. In estimating the response to 

 inoculation two values were noted: (a) the highest titre attained, and 

 (b) the mean titre during the first 50 days after inoculation. The cal- 

 culations were made from the graph drawn from the actual observa- 

 tions. 



The tabulated results show that with a single injection into each 

 of three rabbits of a dose of 10^ bacilli per k.b.w. they attained an 

 immune serum with a highest agglutinating titre of 2550 to 4480 and 



