122 LABORATORY BACTERIOLOGY 
Making the test. Three cc. of the test fluid are placed in 
each of several small test tubes. With a sterile pipette, the 
diluted serum is added to the tubes of test fluid and thoroughly 
mixed. In making the different dilutions, the amount of diluted 
serum to be used is readily ascertained by the following table : 
ae oe Ditton 
I-40 1.2 CC. 3 ce. 1-100 
I-40 0.6 3 1-200 
1-40 0.405 3 1-300 
I-40 0.3 3 1-400 
1-40 0.24 3 I~500 
I-40 0.195 3 1-600 
I-40 0.15 3 1-800 
I-40 0.12 3 I-1000 
1-40 0.105 3 I-1200 
1-40 0.09 3 I-I 500 
I-40 0.06 3 I-2000 
1-40 0.03 3 1-4000 
1-40 0.015 3 1-8000 
Where dilutions greater than 1 to rooo are made, a serum 
diluted 1 to 80 may be used to better advantage, unless the 
pipette employed is very finely graduated. In this case the 
amount of diluted serum for a certain dilution must be double 
that indicated in the table. 
The mixture thus prepared is placed in an incubator at 
37° C. for 24 hours. A temperature higher than 37° C. inter- 
feres with the agglutination. 
' Reaction. The reaction consists of a layer of the aggluti- 
nated bacteria settling and covering the entire convexity at the 
bottom of the tube. This filmlike sediment may become so 
dense that it rolls in at the periphery. The supernatant fluid 
becomes clear in the lower dilutions, but in the higher ones 
the clarification may not be complete, showing that all the 
bacteria have not become agglutinated. This is further 
