524 THE RELATION OF COLLOIDS AND LIPOIDS TO IMMUNITY 



Five constituents enter into the test: 



1. The Antigen. This is an alcoholic extract of syphilitic liver, 

 prepared in exactly the same manner as for performing the Wassermann 

 reaction. High dilutions of the antigen, ranging from 1 : 100 to 1 : 10,000, 

 are prepared with normal salt solution. As in the Wassermann re- 

 action, not every antigen is satisfactory, a point that can be determined 

 only by making preliminary tests. 



2. The patient's serum should be fresh, unheated, and highly diluted, 

 the dilutions ranging from 1 : 100 to 1 : 10,000,000. Usually it is better to 

 use higher than lower dilutions. When too concentrated solutions of 

 serums and of antigen are used, erroneous results are likely to be ob- 

 tained. 



3. A normal solution of sulphuric acid. 



4. A saturated solution of barium hydroxid made equivalent to the 

 normal solution of sulphuric acid. In the use of the barium hydroxid 

 it is imperative to prevent its exposure to the air. A solution that has 

 become cloudy, owing to the entrance of carbon dioxid, should not be 

 used. In carrying out the test it is best to pour out the amount of barium 

 hydroxid needed for the test into a rubber-stoppered bottle or test-tube, 

 so as not to contaminate the stock solution. 



5. A 1 per cent, alcoholic solution of phenolphthalein containing 1 

 per cent, of a 10 per cent, solution of strontium chlorid. The strontium 

 chlorid has been found to catalyze the reaction. 



The Test. The test is conducted as follows: A number of clean 

 beakers of about 50 c.c. capacity are used. For each dilution of the 

 serum a separate beaker is required. One beaker is used for an antigen 

 control, and another to control the system of barium hydroxid and 

 sulphuric acid. Five beakers may be used, Nos. 1, 2, and 3 constituting 

 the main test, No. 4 the antigen control, and No. 5 the system control. 



The reagents are added by means of overflow pipets. To each of the 

 first four beakers is added 1 c.c. of the dilute antigen to be used in the 

 test (about 1 : 10,000). To beaker 5 is added 1 c.c. of the salt solution 

 used in making the dilutions of the antigen and the serums. Now 0.1 

 c.c. of the dilute serum to be tested is added to each of the first three 

 beakers, each beaker, however, containing the same serum in a different 

 dilution. To beaker 5 the same quantity of salt solution is added, but 

 to beaker 4 the antigen control no serum or salt solution is added. 



To each of the five beakers the system of sulphuric acid and barium 

 hydroxid and phenolphthalein is now added carefully. First, 2 c.c. of 

 the normal suphuric acid solution are added to each; then 2 c.c. of the 



