Pleuro-Pneumonia of Cattle. 181 



intervals extending over 5 hours, during which time no alteration 

 was apparent in any of the preparations. Even in Preparation 

 No. 3, which contained Serum No. 30, no agglutination could 

 be recognised under the microscope. 



Agglutination Experiments with Concentrated Culture. 



In conducting the macroscopic agglutination test it had been 

 found that the dilution of culture brought about by the addition 

 •of test serum made it impossible to recognise any opalescence in 

 the fluid in the t tubes. It was therefore considered advisable to 

 .attempt to produce a concentration of the culture so that the 

 .opalescence would be more distinct, and would be clearly visible 

 when the other ingredients of the test were added. Culture 

 in Martin's broth ox serum was placed in centrifuge tubes and 

 whirled in an electric centrifuge at the highest speed the machine 

 was capable of attaining (2500 revolutions per minute) for 4J 

 hours. At the end of that time the tubes were examined, but 

 it was found that it was not possible to effect concentration of 

 micro-organisms in that manner, because the opalescent particles 

 (organisms) in the broth were so fine* and light they could not 

 be thrown down to the bottom of the tubes. 



An attempt was then made to concentrate culture by the eva- 

 poration of some of the fluid medium in which the culture was 

 growing. This evaporation was first tried at room temperature 

 by placing a quantity of culture in a flat dish (Petri dish) inside 

 a desiccator, provided with a circular trough, which contained 

 pure sulphuric acid. Inside the desiccator a more or less com- 

 plete vacuum was established and maintained. It was found that 

 with such an apparatus, the rate of evaporation at room tem- 

 perature was too slow, so the apparatus was placed in the incu- 

 l>ator and evaporation attempted at incubator temperature. At in- 

 cubator temperature, and with the joints of the desiccator sealed 

 with " plasticine " (the temperature made it impossible to use 

 vaseline for the purpose), there was difficulty in maintaining the 

 vacuum, and, altogether, the experiment was not a success. It 

 was then decided to attempt the evaporation of culture at room 

 temperature by using a method, of which the following is a des- 

 cription : — 



424 c.c. of culture in Martin's broth plus ox serum was placed 

 in a flask (Flask A), furnished with a side-arm, and the mouth 

 of the flask was firmly closed with a tightly fitting solid rubber 



