206 PUBLIC HEALTH BACTERIOLOGY 



1. The antigen at first used was a salt solution extract 

 of the liver of a syphilitic fretus. Alcoholic extracts have 

 also been used, and it has been discovered that the 

 extracts of normal liver and spleen, and even a I per cent 

 emulsion of lecithin, will act as antigen. (This shows how 

 little we must know of possible fallacies, in tests like this, 

 with highly complex bodies like liver extract. It does 

 not impugn its specificity for pure antigens, though it 

 suggests the possibility of two different substances, with 

 similar affinities for the immune-body produced by one of 

 them. Of course the probability of this other substance 

 being present, under the conditions of the experiment, is 

 small, but must never be lost sight of.) The quantity of 

 antigen to be used has to be carefully predetermined, as 

 large excess of antigen has been found to cause binding of 

 the complement, even in the absence of the immune-body. 

 A series of trials of varying quantities of antigen with the 

 same amount of complement in each case, shows the 

 largest quantity of antigen which can be used without 

 exerting this action. This amount of antigen is arranged, 

 by proper dilution, to be present in i c.c. 



2. Washed Red Blood Cells. Som sheep's blood is 

 gathered under aseptic precautions into a small sterile 

 flask, containing sterile solution of sodium citrate (0-5 per 

 cent) and sodium chloride (0-85 per cent). The corpuscles 

 are separated by centrifugalizing, and washed repeatedly 

 in the same way with sterile salt solution, to get rid of 

 serum-complement and serum-precipitins. They are then 

 brought down to a 5 per cent emulsion in salt solution, by 

 mixing them with 19 times their bulk of the same. 



3. Hcemolytic Serum for Sheep's Cells. This is obtained 

 by injecting a rabbit with washed red blood cells of a sheep, 

 obtained as above. Of the 5 per cent emulsion, three or 

 four injections are given at intervals of 5 to 6 days ; the 

 first injection of 5 c.c., the second of 10 c.c., the third of 

 15 c.c., and the fourth of 20 c.c. ; intravenously or intra- 

 peritoneally. Ten days later than the final injection the 

 serum is obtained by drawing blood from the carotid 

 artery, allowing it to clot, and pipetting off the serum. 

 A high-potency serum is desirable so that a small quantity 

 only may be required. This obviates or reduces the risk 



