Pleuro- Pneumonia of Cattle. 189* 



that when antigens consisting of pure cultures of the organism, 

 of contagious pleuro-pneumonia in Martin's broth were used,- 

 complement fixation did not occur. 



These results were capable of one of two interpretations : — 



(1) That culture was unsuitable for antigenic purposes; 



(2) That complement fixing antibodies were not present, 

 in the serum of animals affected with contagious pleuro- 

 pneumonia. 



The first interpretation was temporarily accepted as the more 

 probable. The second could not be accepted without further 

 evidence; so, in order to determine whether complement fixing; 

 antibodies were or were not present in the serum of animals 

 affected with contagious pleuro-pneumonia, tissue extracts were, 

 next prepared and tested as antigens. 



The first tissue extracts prepared were saline extracts of 

 diseased lung tissue, taken from animals showing active and ex- 

 tensive lesions of contagious pleuro-pneumonia on post-mortem- 

 examination. These saline extracts of diseased lung tissue were 

 prepared as follows: — 



Carefully selected diseased lung tissue was cut into small pieces- 

 and ground up in a mortar with a little sterile sea sand. Four 

 times its weight of carbol saline solution was added, and the 

 mixture placed in a tightly stoppered bottle, and shaken for five 

 hours in a mechanical shaking apparatus. The mixture was then 

 filtered through gauze to remove all the coarser particles of the 

 tissue. It was then placed in the ice chest for a week, to allow 

 sedimentation to take place, after which time the supernatent. 

 fluid was carefully pipetted off without disturbing the deposit. 

 The supernatent fluid — which is yellowish-brown in colour, and 

 slightly opalescent — was then diluted 1 part in 10 with 0.9 per 

 cent, saline solution, and was used in that dilution in the test, 

 after titration to determine the anticomplementary unit. 



These saline extracts were thoroughly tested with a number of 

 known positive and known negative sera, but in no instance was 

 a positive fixation result obtained. 



A saline extract of the sub-epidermal tumour tissue removed 

 from the inflammatory tumour, which had developed behind the 

 shoulder of an experimentally inoculated animal (Experimental 

 Cow 4) was next prepared. This tissue was not regarded as en- 

 tirely satisfactory for test purposes, for, at the time of its re- 

 moval, the tissue was showing marked evidence of necrosis. The 

 method adopted in the preparation of this tissue extract differed 



