82 ANTIBODIES, IMMUNITY, WASSERMANN SERUM TEST 



bulk. It is now filtered off from the sand, etc., and the clear alcoholic 

 extract, placed in a tightly glass-stoppered bottle, is ready for use at 

 any time. This alcoholic extract of guinea-pig's heart keeps for 

 many months at room temperature, provided it is protected against 

 evaporation and contamination. 



The blood sera used for the tests are the serum from the patient 

 to be examined, the serum from a healthy person and a serum from 

 one who is in the early stages of syphilis. The two latter sera are, 

 of course, used as controls in connection with tests of the patient's 

 serum. Blood is obtained from the three persons with a sterile, all- 

 glass, large hypodermic syringe, the barrel of which will hold from 

 five to ten cubic centimeters of blood. After the latter has been 

 drawn under all aseptic precautions from a vein of the arm, it is at 

 once placed in a centrifuge tube. After coagulation the clot is loosened 

 from the walls of the tube with a platinum wire, the tube is cen- 

 trifuged, and the serum pipetted off. If not clear, it is centrifuged a 

 second time. After the three clear sera have been obtained they are 

 heated for thirty minutes" at 56 C. on a water bath. This inactivating 

 is done to destroy the complement present, but does not interfere with 

 the syphilitic antibody or syphilitic amboceptor. 



8. Everything is now ready to make the test, in the following 

 dilutions. The alcoholic antigen extract is used in a dilution of two 

 in ten, with physiologic salt solution. The rabbit's inactivated serum 

 which contains the hemolytic amboceptor is used in a dilution of one 

 drop in forty or one-quarter of a drop in ten of the salt solution. The 

 reagents are now employed as follows : 



(a) Alcoholic extract of antigen (dilution two in ten). 



(b) Three inactivated sera, one from the patient, two controls. 



(c) The fresh unheated serum from the guinea-pig containing the 

 hemolytic complement. 



(d) The inactivated rabbit's serum (dilution one in forty) contain- 

 ing the hemolytic amboceptor. 



(e) The 5 per cent, washed sheep's corpuscles salt solution suspen- 

 sion, containing the sheep's corpuscles, on which hemolysis will be 

 tried. 



The test is now made as follows: 



Prepare six test-tubes and label as follows : Patient No. 1 ; Patient 

 No. 2; Positive control, No. 1 ; Positive control, No. 2; Negative con- 

 trol, No. 1; Negative control, No. 2. Also label the tubes in this 

 order from No. 1 to No. 6, consecutively. Now add to each tube as 

 follows: * 



To No. 1 (patient No. 1), 10 drops alcoholic antigen extract, plus 

 2 drops of patient's serum, plus 8 drops of salt solution, plus 1 drop 

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



To No. 2 (patient No. 2), 10 drops of salt solution, plus 2 drops of 

 patient's serum, plus 8 drops of salt solution, plus 1 drop of guinea- 

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



