SERUM DIAGNOSIS 475 



The necessary quantity has been established by Schiitz 

 and Schubert as 0.2 and 0.1 cc. placed in two different tubes. 

 In some instances the tube containing 0.2 cc. may show a fixa- 

 tion of the complement, while the tube containing 0.1 cc. of 

 the same serum may show only a partial fixation or hemolysis. 

 In the majority of cases, however, the fixation is usually mani- 

 fested in both tubes.. 



Method of performing test. Four tubes are employed 

 for testing the serum of each animal. Two tubes are used for 

 the test proper and two for controls. The tubes are num- 

 bered 1, 2, 3, and 4. One cc. of a physiological salt solution is 

 added to tubes Nos. 1 and 3. Two cc. is placed in tubes 2 and 

 4. The serum of the suspected horse is then added. This 

 serum has previously been rendered inactive, that is, the com- 

 plement has been "inactivated" by heating for one-half hour 

 at 56-57 C. One-tenth cc. of this inactivated serum is added 

 to tubes 1 and 2. Two-tenths cc. is added to tubes '3 and 

 4. The antigen (glanders bacteria extract) is now added to 

 tubes Nos. 1 and 3. One cc. of an established dilution is used. 

 Tubes 2 and 4 are controls to see whether the suspected horse 's 

 serum will influence hemolysis. 



To each tube is now added 1 cc. of a dilution of the com- 

 plement (blood serum of normal guinea pigs). The proper 

 dilution has been determined by titration as mentioned 

 before. 



A series of six controls should be made in connection with 

 each series of tests carried out. One set of controls will do for 

 a single day's testing as long as the same substances (antigen, 

 complement, amboceptor) are used in each test. It is also 

 well to set what are called "positive" and "negative" con- 

 trols in connection with each series of tests made. That is, 

 the blood from a known glandered animal and likewise the 

 blood from a healthy animal should be tested in connection 

 with the blood from the suspected animals. 



The six controls mentioned above are made according to 

 Table VI beginning with tube No. 5. 



Each tube is shaken carefully and placed in an incubator 



