Blood Fluids in Intracellular Digestion. 



339 



This experiment was repeated again and again, sometimes with slight 

 modifications, such as allowing the serum to act on the bacteria for varying 

 periods before the washed corpuscles were added, but the result was always 

 the same, namely, intra-leucocytic digestion could only be demonstrated when 

 the organisms had been acted on by fresh unheated serum. 



These experiments, although they furnished data showing conclusively 

 that intra-leucocytic digestion only took place in the presence of unheated 

 serum, were unsatisfactory, in that they failed to give any idea as to the 

 proportion of organisms which were digested after being taken up by the 

 leucocytes. The following experiments were therefore undertaken with the 

 view of elucidating this point. 



Phagocytic mixtures consisting of, on the one hand, washed corpuscles, 

 unheated serum, and an emulsion of plague bacilli, and, on the other hand, of 

 washed corpuscles, heated serum, and an emulsion of plague bacilli, were 

 incubated at 37° C. in a water-bath. 



At varying intervals samples of these mixtures were withdrawn and 

 microscopical preparations were made and the number of bacilli contained in 

 100 leucocytes was ascertained. 



In the case of the phagocytic mixture which contained the unheated serum 

 it was found that the number of bacilli that could be recognised in the 

 leucocytes became smaller and smaller the longer the tubes were incubated, 

 at any rate up to a period of four hours, and from these figures it was possible 

 to make a rough estimation of the number of organisms that had been 

 digested. 



In the case of the mixture which contained the heated serum the result 

 was completely different, since each successive sample showed tbat the 

 leucocytes contained larger and larger numbers of bacteria and these appeared 

 normal as regards both their shape and staining reaction. 



The details of one such experiment are here given. 



A very thick emulsion of plague bacilli, from a 24-bour-old agar culture, 

 was made in normal saline to which 1 per cent, of formalin had been added. 

 The emulsion was kept at room temperature for about one hour to allow the 

 formalin to act on the bacilli. 



Tins procedure, which killed the organisms, was necessary, as when similar 

 experiments were made with emulsions of living plague bacilli it was found 

 that after incubation at 37° CL for some hours a proportion of the bacilli 

 which had been ingested by the leucocytes were capable of multiplying, and 

 in consequence of this the results obtained were irregular and fallacious. 



This thick emulsion was now diluted 100-fold with normal saline, so that 

 the concentration of formalin in the final phagocytic mixture given below was 



