AYASSERMANN AND OTHER COMPLEMENT FIXATION TESTS 245 



For each serum that is to be subjected to the Wassermann test three tubes 

 are used; one of these tubes during the first hour of incubation contains the sus- 

 pected patient's serum and guinea-pig serum only, no syphilitic antigen ; this is 

 a control for the patient's serum. Only sera which contain nothing that can 

 inhibit hemolysis, other than syphilitic amboceptors, are susceptible to exami- 

 nation by the complement fixation test. Since amboceptor cannot fix comple- 

 ment except in the presence of antigen, and there is no antigen in this tube, then 

 the complement must remain free and unaltered, regardless of whether the serum 

 is syphilitic or not. 



If the complement in this tube is fixed or vitiated during the first hour of 

 incubation it is evidence that the human serum contains something, not relative 

 to syphilis, which makes impossible its examination. 



In other words, the tube that contains no syphilitic antigen must always 

 show complete hemolysis at the end of a Wassermann test; if it does not there 

 is no diagnostic significance to the findings. 



The writer has never seen a human serum cause this phenomenon when 

 subjected to examination within 3 days after withdrawal from the circulation, 

 but human sera, even when sterile, occasionally do so to a slight degree several 

 weeks after they have been obtained from a patient; putrid sera nearly always 

 vitiate complement. This control is therefore indispensable. 



TECHNIQUE OF THE WASSERMANN TEST 



1. Bleed patient or patients. 



2. Compute the number of tubes that will be used in making the tests; 

 allowing o.i cc. of guinea-pig serum for each tube, compute the amount required. 



3. Obtain sufficient guinea-pig blood to yield 10 per cent, more than com- 

 puted amount of serum required. 



4. Collect blood in citrate solution to furnish red cells. 



5. Wash and make a 5 per cent, suspension of the red cells in normal salt 

 solution. 



6. Make a 10 per cent, solution of the guinea-pig serum. 



7. Make a i per cent, solution of sensitized rabbit serum. 



8. Titrate or standardize the complement. 



9. Separate patient's serum from clot and inactivate it. 



10. Compute the amount of antigen required and mix the alcoholic extract 

 with normal salt solution. 



11. Place three tubes in the rack for each patient's serum to be tested, also 

 three for the positive control serum and three for the negative control serum, 

 place three tubes at the extreme right for complement control, hemolytic sys- 

 tem control, and antigen control (see Fig. i). 



12. Put one unit of antigen in every tube in the middle and bottom rows 

 except the hemolytic system control tube and the antigen control tube, put no 

 antigen in the hemolytic system control tube, put two units of antigen in the 

 antigen control tube, do not put antigen in any tube in the top row. 



13. Put o.i cc. of patient's serum in each of the three tubes provided for it, 



