198 G. G. Heslop: 



plement (M.H.D.) is ascertained by titrating the complement 

 with red blood cells, sensitised by more than a single unit of 

 amboceptor, it is possible to arrive at a unit of complement, which 

 is just large enough to effect complete haemolysis of the sensi- 

 tised red blood cells, in the case of a negative serum, but which 

 at the same time is small enough to be more or less completely 

 fixed by the antigen- — antibody combination in the case of a. 

 positive serum. 



This " overloading " of the haemolytic system with ambo- 

 ceptor cannot be increased beyond certain limits. While a slight 

 increase of the amboceptor factor is a decided advantage, the 

 increase must not exceed two complete units, for it has been 

 found that, owing to the feeble nature of the fixation which, 

 occurs in contagious pleuro-pneumonia, a large increase in the 

 amount of amboceptor in the haemolytic system tends to disso- 

 ciate some of the complement already held by the antigen-posi- 

 tive serum combination. In consequence of this, a large excess 

 of amboceptor only tends to shorten the time required for haemo- 

 lysis to occur. 



It has been found that the best results have been obtained by 

 using 1^ units of amboceptor to sensitise the red blood cells, and 

 then to titre the complement against one unit of these over sen- 

 sitised cells, in order to determine the absolute M.H.D. of com- 

 plement required for haemolysis. 



Technique of the Complement Fixation Test for 

 Contagious Pleuro-Pneumonia. 



The method of carrying out the test is as follows: — ■ 

 0.5 c.c. of a 5 per cent, suspension of sheep's red blood cells- 

 equals 1 unit. Against this unit, the haemolytic amboceptor is 

 titrated, and it is found that 0.5 c.c. of 1 in 1000 dilution equals 

 one unit of haemolytic amboceptor. Each unit of red blood cells 

 is sensitised by 1J units of haemolytic amboceptor, by mixing 

 together the necessary units of amboceptor and red blood cell 

 suspension, and allowing them to stand for one hour. The mix- 

 ture is then centrifuged, and the clear fluid pipetted off. The 

 sensitised red blood cells are then washed with saline solution,, 

 and are resuspended in saline in order to make a 5 per cent, sus- 

 pension. 0.5 c.c. sensitised red blood cells (5 per cent, suspen- 

 sion) equals one unit. 



The complement (guinea-pig's serum, 1 in 10), is now titrated 

 against one unit of sensitised red blood cells, by testing it in; 



