E. A. Watson 
167 
Tube No. 1 will show only slight or partial haemolysis; as one 
descends the series the reaction is seen to be increased, until, usually 
between Nos. 4 and 7, a tube is reached in which the reaction is abso¬ 
lutely complete. The first tube in the series in which all the red cells 
are completely dissolved indicates the complement titre. If this 
occurs in tube No. 5, for example, then 0-55 c.c. of a 1: 20 dilution of 
complement is the titre, equivalent to TO c.c. of a 2-75 per cent, dilution. 
For the antigen titration and final tests the complement is accordingly 
made up so that TOc.c. of the dilution contains the amount of complement 
indicated by the above titration. 
From now on it is optional whether one employs the relatively 
large amounts of reagents as given in the first set of complement titration, 
or the one-half amounts as in the second set. The latter is the more 
economical, especially when a large number of tests are being performed, 
and is given personal preference to by the writer as it seems to provide 
an even more highly sensitive test reaction than when the larger amounts 
are employed. 
(3) Titration of Antigen. 
Dilute 1-0 c.c. of stock trypanosome antigen with 19-0 c.c. of normal 
salt solution. 
Prepare the complement and haemolytic serum according to their 
titration values already determined. 
Inactivate by heating for half-an-hour at 58° C. in a water bath, 
2-0 c.c. of known positive dourine horse serum and 2-0 c.c. of known 
negative or normal horse serum. 
The antigen is then titrated in a double set, the one being with the 
positive serum, the other with double the amount of negative serum. 
Thus: 
Positive set Negative set 
Controls 
Salt sol. 
C.C. 
Known 
positive 
horse 
serum 
c.c. 
Antigen 
c.c. 
Comple¬ 
ment 
c.c. 
Salt sol. 
C.C. 
Known 
negative 
horse 
serum 
c.c. 
Antigen 
c.c. 
Comple¬ 
ment 
C.C. 
Tube No. 1 
1-0 
0-1 
0'02 
0-5 
1-0 
0-2 
0-05 
0-5 
99 
2 
1-0 
0-1 
0-05 
0*5 
1-0 
0-2 
0-1 
0-5 
19 
3 
1-0 
0-1 
0-1 
0-5 
1-0 
0-2 
0-2 
0-5 
19 
4 
1-0 
0-1 
0-15 
0-5 
1-0 
0-2 
0-3 
0-5 
:i 
5 
1-0 
0-1 
0-2 
0-5 
1-0 
0-2 
0-4 
0-5 
9 
6 
1-0 
0-1 
0-25 
0-5 
1-0 
0-2 
0-5 
0-5 
'.9 
7 
1-0 
0-1 
0-1 
0-5 
1-0 
0-2 
0-6 
0-5 
99 
8 
1-0 
0-2 
— 
0-5 
1-0 
0-2 
— 
0-5 
19 
9 
1-0 
— 
0-25 
0-5 
1-0 
— 
0-5 
0-5 
99 
10 
1-5 
—- 
— 
0-5 
2-0 
— 
— 
— 
11—2 
