78 DISEASES OF DOMESTICATED BIRDS 
in the quantity of 1 cc. to each tube. A series of tubes is em- 
ployed for each serum, depending on the number of serum dilu- 
tions to be made. In practice, a serum which causes agglutination 
in a dilution of .01 or even .02 is sufficient to establish a positive 
diagnosis. Using 1 ¢.c. as the standard amount of bacterial sus- 
pension, sera which have been diluted by the addition of normal salt 
solution to the extent of .04, .02, and .01 per cent of the original, 
are distributed in their respective tubes in quantities of 1 c.c. each. 
The tubes are labeled to show the identity of the various sera and 
the dilutions. In practice, it is found convenient to dilute the serum 
to 10 per cent of its original volume and add .4, .2, and .1 of a ce. 
to the 1 c.c. of bacterial suspension. The tubes are shaken to insure 
a thorough mixture of the contents and may be set aside at a tem- 
perature of 37° C. for from twenty-four to forty-eight hours, when 
readings may be made. Instead, when rapid diagnosis is desired, 
they may be placed in an incubator at 87° C. for 4% hour and then 
centrifuged at the rate of 1500 to 1800 revolutions to the minute for 
several minutes. Either course gives similar results. In positive 
cases the organisms in the fluid collect in flocculent masses which 
have a tendency to form a diffuse coating over the bottom of the tube, 
while in the negative cases the organisms are deposited in a small 
circular mass in the center of the bottom of the tube. This is par- 
ticularly apparent where centrifugalization has been employed. On 
shaking the tubes the clearly positive ones will show a clear fluid 
containing flocculent masses while the negative ones are uniformly 
clouded. 
The agglutinating serum dilution which will establish a diagnosis 
has not been definitely fixed. It is a well known fact that normal 
blood serum is capable of agglutinating organisms to a greater or 
less degree. In B. pullorum infection tests, a serum which in the 
amount of .02 c.c. or less causes agglutination, undoubtedly indicates 
a positive case. 
Intradermal test. This test depends for its results upon the 
principle underlying allergic reactions. It is very similar in char- 
acter to the intradermal tuberculin test, especially as used to detect 
tuberculosis in fowls. 
A culture of several strains of B. pullorum from widely separated 
sources is grown in bouillon at a temperature of 37.5° ©. for a 
period of thirty days to three months. Whether these time limits 
are necessary has not been determined. The product is then steril- 
ized at a temperature of 60° C. for a period of one hour and carbolic 
