1904.] Combining Properties of Serum-Complements, etc. 3 



: 



combination with R + IB can be dissociated, but it will be well to give 

 a detailed experiment in illustration. The red corpuscles of the ox 

 turated with the corresponding IB obtained from the rabbit can take 

 about 8 9 hsemolytic doses of guinea-pig's C, and if we make the 

 uspension of red corpuscles four times the usual strength, i.e., 20 per 

 cent., then 1 c.c. of such a suspension will take up more than 30 doses 

 of C. We can then test whether any of the C is recoverable. The 

 following are the details : 



1. 1 c.c. of a 20-per-cent. suspension of ox's corpuscles in salt solution is placed 

 in a test-tube, and there are added (a) more than sufficient IB to saturate the 

 corpuscles, (b) 1 c.c. of guinea-pig's C (the M.H.D. is 0'055 c.c.), so that nearly 

 twenty doses are added. 



2. The tube is placed in the incubator for 2 hours at 37 C., the contents being 

 well shaken from time to time. 



3. At the end of 2 hours 0'5 c.c. of a 5-per-cent. suspension of ox corpuscles, 

 treated with IB, is added (the hsemolytic dose of C for this is, of course, 0'027 c.o.) ; 

 the tube is placed in the incubator for another hour. 



4. The contents of the tube are then centrifugalised, and the corpuscles are 

 washed with salt solution to remove the red fluid. The corpuscles are then laked 

 with water, and compared with 0'5 c.c. suspension of red corpuscles similarly 

 treated. 



The result is that no perceptible haemolysis of the added corpuscles 

 has taken place. By the method employed the haemolysis of less than 

 a fifth of the corpuscles is readily detected, i.e. t if 0*005 c.c. of C were 

 recoverable, this would be shown by the resulting haemolysis. Such 

 an experiment, therefore, shows (a) that practically complete com- 

 bination of the added complement takes place, and (b) that of the 

 combined complement not 1 per cent, can be dissociated. 



When the saturation point has been reached, i.e., when C is first 

 obtainable, any C then added remains free to a large extent. In a 

 former paper it was stated that, as a rule, a single dose of C added 

 remains free, but further experiments show that this is rather the 

 exception. It sometimes occurs, but usually after the saturation point 

 more than one dose of C requires to be added to give one dose free. 

 Considerable variations occur in this respect, and we are unable to 

 explain them, though possibly the presence of a certain proportion of 

 complementoid mixed with the complement may be the cause. This, 

 however, does not affect the statement regarding the firmness of the 

 union of with R + IB after combination has taken place. 



When we come to the combination of C with anti-C, we find that 

 quite another condition obtains. Here we have an exaggerated 

 instance of what Ehrlich found in the case of a toxin and anti-toxin, 

 viz., that when a neutral mixture of these substances is made, and 

 time is allowed for combination, the addition of several lethal doses of 

 toxin may be necessary before a lethal dose remains free. The 

 following may be taken as examples : 



B 2 



