136 JOHNE'S DISEASE 



culture of the tubercle bacillus as recommended by 

 Arloing. Owing to the slow growth of Johne's 

 bacillus in the substance of a fluid medium, even 

 though it contains a certain amount of an alcoholic 

 extract of B, phleij one is obliged to have recourse 

 to an emulsion of the bacilli grown on solid media. 

 As will be remembered, Koch and Romberg used an 

 emulsion made from a growth on solid medium in their 

 tubercle agglutination reactions. To obtain an emul- 

 sion of Johne's bacillus, the growth is mixed with 

 saline, placed in a stout bottle containing glass beads» 

 and shaken in an ordinary electric shaker. To obtain 

 a good homogeneous emulsion, the usual precautions 

 must be taken. The culture should be young, and 

 after shaking should be filtered or centrifuged, to free 

 the emulsion from large clumps of bacilli. Fresh 

 unheated serum should be used, and it is best to per- 

 form the test as soon as possible after the withdrawal 

 of the blood. This precaution does not refer to fowl 

 serum, which agglutinates as well after heating at 56° C. 

 as before. In this case, too, the serum may be kept in 

 an ice-chest for months without any apparent diminu- 

 tion in activity. As in testing a tubercular serum, the 

 mixture must be incubated for some time — two to 

 four hours. If the microscopical method of examina- 

 tion is used, two hours is usually sufficient, and this 

 method in our opinion is far more reliable than the 

 macroscopic test. A control with the fresh unheated 

 serum of a normal animal of the same species as that 

 from which the suspected serum was obtained must, 

 of course, be made. It is well also to have a second 

 control, containing no serum, as spontaneous agglu- 

 tination of the emulsion is liable to take place, especi- 

 ally if living bacilli are used. 



The sera of seven naturally infected cattle, six of 



