376 THE ALCALIGENES— DYSENTERY— TYPHOID GROUP 
from day to day in the titer of serums from typhoid cases. Controls 
must always be made. The typhoid culture is diluted with an equal 
volume of salt solution. Spontaneous agglutination sometimes takes 
place when no serum is present. This is shown in the control and at 
once invalidates the agglutination which may be obtained with the 
serum. 
Techniqve of Test. — (A) Microscopic Method.— Dried blood, blood 
serum, blister fluid, or whole blood is diluted 1 to 20 with physio- 
logical salt solution. A loopful of this diluted fluid is mixed intimately 
with a loopful of typhoid broth c^ilture on a coverglass and suspended 
in a hanging drop slide ringed with vaseline to prevent evaporation. 
The final dilution of the blood is 1 to 40 by this procedure. A control 
is made using a loopful of salt solution and a loopful of typhoid culture 
prepared in the same manner. Both the serum and the control are 
kept at room temperature. A preliminary examination should show 
actively motile bacteria in the control preparation and usually actively 
motile bacteria in the serum preparation. It sometimes happens that 
agglutination takes place in the serum preparation almost immediately. 
If the preliminary examination is satisfactory, the final examination 
is made at the end of an hour. Both preparations are examined and 
the controls should show actively motile unclumped organisms. A 
positive agglutination is recorded if the control is as stated and the 
organisms in the serum preparation are non-motile and gathered 
together in clumps with few or no free-swimming bacteria between 
the clumps. 
(B) Macroscopie Method.— Yarious dilutions of serum are placed 
in small sterile test-tubes, 1 cc. in each test-tube. As a routine, a 
dilution of 1 to 20 is used, but a series of dilutions up to the limits 
of the serum are frequently made. To each tube is added 1 cc. of a 
broth culture of typhoid bacilli. A control is made by adding 1 cc. 
of a broth culture of typhoid bacilli to 1 cc. of salt solution. These 
mixtures are respectively shaken and incubated together with the 
control at 37° C. for two hours and examined; then they are placed in 
the ice-box, and examined once again eighteen to twenty-four hours 
later. A positive agglutination is indicated when the supernatant 
fluid of the serum typhoid mixtures is clear, while the control containing 
no serum remains uniformly cloudy. 
The microscopic method is much more rapid than the macroscopic 
method and is sufficiently accurate for ordinary purposes. The macro- 
scopic method requires a longer time, but it is more accurate, for the 
dilutions can be made carefully with graduated pipettes. 
Discussion.— Axa'ilahh statistics show that about 20 per cent of 
typhoid patients exhibit a positiAe agglutination reaction at the end 
of the first week; ()0 per cent at the end of the second week; 80 per cent 
at the end of the third week; and 90 per cent at the end of the fourth 
week. These agglutinins persist; about 75 per cent of all patients 
exhibit a positive agglutination after two months. Occasionally 
