440 THE TECHNIC OF COMPLEMENT-FIXATION REACTIONS 



As the complement is " fixed" by the syphilis antibody and ex- 

 tract, hemolysis could not occur when the corpuscles and amboceptor 

 were added. If a portion of the complement is fixed by antibody and 

 extract, then the unfixed portion will hemolyze some of the corpuscles, 

 the reaction being moderately positive, slightly positive, etc., depend- 

 ing upon the degree of hemolysis that takes place. This illustrates 

 the importance of observing exactness in pipeting, and the great 

 influence of quantitative factors in testing for the Wassermann reac- 

 tion, for if an excess of complement is used, there may be sufficient 

 for all the syphilis antibody, and enough unbound complement to 

 hemolyze all the corpuscles. In this manner a false negative reaction 

 will result. Corpuscles and sufficient hemolytic amboceptor are 

 added merely in order to test for any free complement. Under proper 

 conditions a total lack of hemolysis indicates that there is no free 

 complement, but that it has been fixed by syphilis antibody and 

 extract, constituting a positive reaction (+ + + +). Complete 

 hemolysis indicates that complement was not bound and that syphilis 

 antibody was, therefore, absent from the fluid tested a negative 

 reaction ( ). Partial hemolysis indicates that a portion of the 

 complement has been fixed by smaller amounts of syphilis antibody 

 and of the extract, yielding partially positive reactions (+ + +; 



+ +; +; *). 



6. The front tube containing the known normal serum should show 

 complete hemolysis because, in the absence of syphilis antibody, the 

 complement remains free to hemolyze the corpuscles with the hemo- 

 lytic amboceptor. 



7. Various methods have been proposed for recording the results 

 of hemolytic tests. The following scheme, after Citron, is widely 

 used (Fig. Ill): 



+ + + + = complete inhibition of hemolysis = strongly positive. 

 + + + = 75 per cent, inhibition of hemolysis = moderately positive. 

 + + = 50 per cent, inhibition of hemolysis = weakly positive. 

 -f- = 25 per cent, inhibition of hemolysis = very weakly positive. 

 =*= = less than 25 per cent, inhibition of hemolysis = delayed 



hemolysis or doubtful reaction. 

 = complete hemolysis = negative reaction. 



Under the third method a scale is given that is easily prepared 

 for making these readings. However, after some experience they are 

 readily made, and at first should be attempted only after the non- 

 hemolyzed corpuscles have been centrifuged or allowed to settle to the 



