128 METHODS OF EXAMINING SEEUM 



and Levaditi, however, found that an extract of normal guinea- 

 pig's liver along with syphilitic serum fixed complement, i.e., 

 acted as antigen, and subsequent observations showed that ex- 

 tracts of other tissues are also more or less efficient, as are also 

 certain definite substances, such as sodium oleate, sodium glyco- 

 cholate, lecithin, mixtures of such, and especially mixtures of 

 lecithin and cholesterin, etc. Although abundant observations 

 have established the validity of the test as a means of diagnosis, 

 the reaction which led to its discovery is no longer sufficient to 

 explain it, and its nature is not yet understood. 



In order to carry out the test, we require (a) serum from the 

 suspected case, (b) an extract of liver or other organ, (c) the fresh 

 serum of an animal to act as complement, and (d) sensitised ox 

 corpuscles, i.e., a 5 per cent, suspension of washed ox corpuscles to 

 which several doses of immune-body have been added (p. 126). 



The following may be given as an example of the method as 

 formerly used : — 



Add to a small test-tube containing 0'5 c.c. of 0'8 per cent, sodium 

 chloride solution :— (a) 0'05 c.c. of serum from the suspected case (heated 

 for an hour at 55° C. to destroy the complement), (6) O'l c.c. of an extract 

 of guinea-pig's or ox's liver, and (c) a certain amount, usually O'l c.c. 

 of guinea-pig fresh serum to act as complement. Place in incubator at 

 37° C. for an hour and a half. At the end of that time add 1 c.c. of sen- 

 sitised ox corpuscles and place in the incubator for another hour. If at 

 the end of the hour the corpuscles are not lysed the complement has been 

 fixed in the first stage — the result is positive as regai-ds the presence of 

 syphilis ; if lysis has occurred the result is negative. Controls were used to 

 test effect on complement of {a) suspected alone and (6) antigen alone. 



It is to be noted, however, that the substance in the syphilitic 

 serum which leads to the fixation of complement varies greatly 

 in amount in different cases, and it is not possible to state abso- 

 lutely the quantity of complement which must be fixed in order 

 to give a positive result. Manifestly there will be cases where 

 the amount fixed is just under any standard adopted, and these, 

 which are to be regarded as suspicious or doubtful, will be missed 

 with a one-tube method. This is well exemplified in cases under- 

 going treatment with salvarsan. Moreover, the amount of com- 

 plement, as estimated by the hsemolytic dose, varies considerably 

 in different samples of fresh serum. It is accordingly necessary 

 for satisfactory results to estimate the hsemolytic dose of the 

 guinea-pig's serum, and to prepare a series of tubes, each con- 

 taining the same amounts of serum and of antigen, but with a 

 different number of doses of complement in each tube. In this 

 way we can find the number of doses of complement deviated 

 in each case. As controls, the effect on doses of complement 



