BACTERIAL COMPLEMENT FIXATION TECHNIQUE 409 



The test dose based upon using one-third of this would be 0.5 

 c.c. of a 1 :6 dilution of antigen. If, instead of the above, antigenic 

 titration was done and it was desired to use 10 antigenic units in 

 the test, it would only be necessary to use 0.5 c.c. of a 1 :20 dilution 

 of this hypothetical antigen since one antigenic unit is 0.5 c.c. of 

 1 :200 dilution. 



After the antigenic titration has been completed and a test 

 dose of antigen is decided upon, one is then ready to set up a com- 

 plement fixation test to determine whether antibodies correspond- 

 ing to the antigen are present in the patient's serum. In sucli a 

 test it is necessary to establish by experiment (adequate controls) 

 the validity of the results obtained. Three methods are available 

 for complement fixation. One is the quantitative, a second is the 

 qualitative, and third is the simplified test. A protocol for the 

 simplified test is given in Table XI and is illustrated in Plate VII. 

 In order for the student better to understand the significance of 

 the results indicated in Table XI and Plate VII the following 

 discussion is presented : 



From a careful inspection of tlie above table it will be evident 

 that Tube 5, which contains patient's serum, antigen, complement 

 and sensitized red cells, constitutes the only test of the patient's 

 serum for the presence of complement fixing antibodies. The other 

 nine tubes are obviously controls. These controls are not only 

 necessary from the standpoint of accuracy of results, but also fre- 

 quently enable the laboratory worker to ascertain the cause of his 

 failure. 



No Hemolysis in Tube 5. — If either no hemolysis or partial 

 hemolysis occurs in Tube 5 containing patient's serum, antigen, 

 complement, red cells and hemolysin, the following questions arise : 



1. Is it because the patient's serum contains sufficient antibodies 

 to sensitize the antigen and thus give specific fixation? 



2. Is the patient's serum anticomplementary? 



3. Is the antigen anticomplementary? 



4. Is it because of insufficient amount of complement due to a 

 drop in titer or to errors in making up the standard dilution or to 

 pipetting into the tubes? 



5. Is it due to hemolysin deficiency for similar reasons ? 



