174 PRACTICAL METHODS IN IMMUNITY 



It is usual to expect strong fixation with a paresis fluid in the smallest 

 amount noted above. For tabes use more fluid, as 0.5 and i c.c. 



NOGUCHI'S METHOD 



For the suspension of red cells use a J^% suspension of washed 

 human red cells. 



For complement use fresh guinea-pig serum in a dilution of i part 

 to i> parts of salt solution (40%). 



Method. Take four small test-tubes (12 by 125 mm.) label la, ib and 20, 2&, re- 

 spectively. Into la and ib each put i drop of the serum of the patient to be tested 

 and into ia and 26 each put i equal size drop of the serum of a person known to give a 

 positive test for syphilis. The small drop delivered by a very finely drawn-out 

 capillary pipette, held vertically, is equal to about 0.02 c.c. or 50 drops to i c.c. 

 Next add to each of all four tubes i c.c. of the ^ % suspension of washed red cells. 

 Then add to each tube o.i c.c. of the 40% fresh guinea-pig serum. Now add 

 to tube ia and tube 2a each o.i c.c. of the i to 10 antigen dilution (opalescent working 

 antigen emulsion). Tubes ib and zb are controls not containing antigen. Mix con- 

 tents of tubes thoroughly and incubate at 37C. for one hour or for one-half hour 

 in a water-bath. Now add to each of the four tubes 2 units of the immune haemolytic 

 serum, as measured off on the amboceptor paper strip thus with a paper of which 2 

 mm. equals i unit, drop into each tube 4 mm. of the strip. 



The tubes without antigen (ib and 2 b) should show good haemolysis. 

 Tube 2d, that of the known syphilitic, with antigen, should not show 

 haemolysis and that of the person examined (ia) should show haemolysis 

 in case the test is negative for syphilis. Moderately positive cases may 

 show a slight trace of haemolysis. Where the tubes without antigen 

 are without colour (no haemolysis) it shows that there is some anti- 

 complementary factor at work and that the tests should be regarded 

 as unsatisfactory. 



This failure to haemolyze in the control tubes may be due to anticomplementary 

 substances in the serum or to the bacterial contamination of the serum, the proteids 

 of the disintegrating bacteria possibly preventing haemolysis by antigenic action and 

 thus absorbing the complement, which otherwise would bring about haemolysis. 



It is advisable also to employ the serum of a person known to be 

 free from syphilis. In this case we should use two additional tubes, 

 30 and 36, conducting the test as for the syphilitic control serum. 



If a normal serum is not used then use a system control in which there are put all the 

 reagents noted above except the human sera from the patient or the syphilitic 

 control. 



When examining a large number of sera at the same time it is well to use rubber 



