Contagious Fleuro-Pneuviouia of Cattle. 185 



Antigen ii. — An alcoholic extract of fresh subepidermal 

 tumour tissue from experimental Calf 10 (Antigen 6) to which 

 was added 0.4 per cent, chlosterin. 



Antigen I2. — An alcoholic extract of dried subepidermal 

 tumour tissue from Calf 10, to which was added 0.4 per cent, 

 chlosterin. 



The chlosterin used in the preparation of Antigens 7, 8, 9, 10^ 

 11 and 12 was a sample freshly prepared in the Physiological 

 laboratories of the Melbourne University. The extracts to which 

 chlosterin was added were found to be so highly anticomplemen- 

 tary that they were useless for testing purposes. Antigens 11 

 and 12 were again made up, but with only half the quantity of 

 chlosterin added, i.e., 0.2 per cent. They were found on testing 

 to be still too anticomplementary for use in a complement fixa- 

 tion test. 



Agglutination Tests. 



A number of blood samples were obtained from animals which 

 were found to be infected with contagious pleuro-pneumonia at 

 the time of slaughter and post-mortem examination. Blood 

 samples were also obtained from a number of animals who were 

 known not to be affected, or to have been in contact with the 

 disease. In this manner a large number of *' known positive '^ 

 and " known negative " serum samples have been acquired, and 

 these sera have been used as test sera in the agglutination tests. 



Microscopic Agglutination. 



Agglutination tests with known positive and known negative 

 sera, using the microscopic method with dark ground illumina- 

 tion, have been carried out with very unsatisfactory results. With 

 a known positive serum, agglutination can be observed invari- 

 ably in a dilution of 1 in 20 in about three and a-half hours 

 after mixing. Unfortunately, however, the majority of knowni 

 negative sera tested also showed agglutination in the same dilu- 

 tion in the same time. In dilutions of 1 in 30, only five out of 

 eight positive sera tested showed agglutination, while two out of 

 8 negative sera tested also showed agglutination in the same 

 dilution. In dilutions of 1 in 35, two positive sera out of eight 

 tested showed agglutination, while none of the eight negative sera 

 tested showed agglutination in that dilution. No serum, either 

 positive or negative, showed agglutination in a dilution of 1 

 in 40. 



