THE WASSERMANN TEST 83 



To No. 3 (positive control No. 1), 10 drops of alcoholic antigen 

 extract, plus 2 drops of serum of syphilitic, plus 8 drops of salt solu- 

 tion, plus 1 drop of guinea-pig's serum, plus 9 drops of salt solution. 



To No. 4 (positive control No. 2), 10 drops of salt solution, plus 

 2 drops of syphilitic serum, plus 8 drops of salt solution, plus 1 drop 

 of guinea-pig's serum, plus 9 drops of salt solution. 



To No. 5 (negative control No. 1), 10 drops alcoholic antigen 

 extract, plus 2 drops of healthy person's serum, plus 8 drops of salt 

 solution, plus 1 drop of guinea-pig's serum, plus 9 drops of salt 

 solution. 



To No. 6 (negative control No. 2), 10 drops of salt solution, plus 

 2 drops of healthy persons serum, plus 8 drops of salt solution, 

 plus 1 drop of guinea-pig's serum, plus 9 drops of salt solution. 



We have now three sets of tubes, each set containing the serum of 

 a different person, and in each set one tube with alcoholic antigen 

 extract and one set without this extract, its place being taken by 

 physiologic salt solution. There must be another set of controls pre- 

 pared from the sera to find out whether the extract is of the proper 

 strength. In these controls the alcoholic extract is present in the fol- 

 lowing proportions : 1 to 10, 2 to 10, 4 to 10, 6 to 10. The extract should 

 be of such strength that 1 to 10 is strong enough to inhibit hemolysis 

 with a serum from a case of syphilis; 4 to 10 should be of such a strength 

 that the presence of a syphilitic antibody is necessary to prevent 

 haemolysis, and 6 to 10 should be strong enough to prevent it alone. 

 These additional tests need not be made every time, but just often 

 enough, so that the alcoholic antigen extract is always kept under 

 control. Another point about this extract is the following: Since 

 it contains in the proper dilution about 25 per cent, alcohol, its 

 drops are smaller than the drops of the purely watery fluids used, 

 hence it is necessary to use a special dropper for the alcoholic extract 

 which has been tested out, or if the same dropper or pipette is used, 

 it is necessary to take from twelve to thirteen drops of the dilute 

 alcohoic extract for each ten drops of the watery solutions. All 

 reagents must be used so that they represent in the salt solutions 

 equal amounts, namely, ten drops. 



After the test-tubes have been prepared they are placed in the 

 incubator for thirty minutes. If there is present in any test-tube a 

 combination of antigen and syphilitic antibody, these will, during 

 the half-hour in the incubator, unite and attract and fix to themselves 

 the complement which is present in each tube. 



After thirty minutes the six test-tubes and the other controls which 

 may have been made are removed from the incubator and to each 

 one is added : 



10 drops of the diluted inactivated rabbit's blood serum contain- 

 ing the hemolytic amboceptor. 



10 drops of the 5 per cent, washed sheep's corpuscles salt solu- 

 tion suspension. 



