THE TECHNIQUE OF SERUM REACTIONS 271 



Tube 1. 1 drop patient's serum + complement (.1 c.c. of 40 per cent guinea- 

 pig serum + antigen. 



Tube 2. 1 drop patient's serum + complement (no antigen). 

 Tube 3. 1 drop known syphilitic serum + complement + antigen. 

 Tube 4. 1 drop known syphilitic serum + complement (no antigen). 

 Tube 5. 1 drop known normal serum + coniplement + antigen. 

 Tvbe 6. 1 drop known normal serum + complement (no antigen). 

 Tube 7. Complement alone (for hemolytic system control). 



To each tube then add 1 c.c. of the one per cent emulsion of human 

 corpuscles. Shake mixtures thoroughly and incubate or place in water 

 bath at 38-40° C. for one hour. Then add to each tube two units of 

 amboceptor and replace in water bath for one hour. At the end of this 

 time in a positive test there will be no hemolysis in tubes one and three 

 while all the other tubes will show hemolysis. 



Noguchi has simplified the technique of complement fixation 

 further by drying measured amounts of antigen and amboceptor upon 

 small squares of blotting paper. These may be dropped into the tubes 

 directly, obviating the necessity of preparing fresh dilutions of the con- 

 centrated substances for each test. The substances in the dried state, 

 moreover, may be preserved for longer periods than when kept in the 

 liquid form. 



The Determination of Antigen by Complement Fixation. — The prin- 

 ciples underlying the preceding tests for the determination of sus- 

 pected antibodies may be equally applied to the determination of 

 suspected antigen. In the former case it was necessary to bring the 

 serum to be tested into contact with the antigen specific for the suspected 

 antibody, in the presence of complement, and at a suitable tempera- 

 ture. At the end of an hour the mixture was tested for free comple 

 ment by the addition of hemolytic amboceptor and red blood cells. 

 In testing for antigen, the procedure is reversed, in that the serum or 

 other substance (bacterial extract) to be tested is brought into contact 

 with an antibody specific for the antigen, in the presence of complement ; 

 and at the end of an hour at suitable temperature, free complement 

 is again determined by hemolytic reaction as before. 



When dealing with bacterial antigen, it is necessary, therefore, to 

 prepare a highly potent immune serum against the bacteria which 

 contain the specific antigen which is sought. 



Thus in testing for typhoid-bacillus antigen in the serum of a patient, 

 the substances required are as follows : 



1. Complement: obtained from fresh guinea-pig serum. It is best 

 to titrate the complement when possible, using for the test double the 



