360 Dr. J, O..W. Barratt, [ Dec. 5, 
Kxperiments in which the Complement Enuployed was that Present in the Serum 
of the Rabbit or of Man. 
Method.—The hemolytic system selected consisted (1) of the red blood cells 
of the rabbit, employed in a 24-per-cent. suspension in 0°85-per-cent. sodium 
chloride solution; (2) of immune body derived from the serum of the rat; 
and (3) of complement contained in human blood serum or that of 
the rabbit. The rat providing the immune serum had been previously 
inoculated intraperitoneally on three occasions at intervals of 10 to 14 days, 
receiving each time about 0°3 gramme of the red blood cells of the rabbit, 
previously washed in 0°85-per-cent. solution of sodium chloride. Before being 
used the serum was inactivated by heating to 57° C. for one hour. The 
hemolytic power of the serum of the rat varied with different animals; that 
used in the experiments recorded in the tables was capable of completely 
heemolysing, in one and a-half hours, at a temperature of 37° C., four to five 
tines its volume of the red blood suspension employed. In sensibilising the 
red blood cells of the rabbit one part of serum was added to one part of 
the red blood cell suspension, and the mixture allowed to stand at 37° C. 
with occasional shaking, for one hour and a-half. A slight degree of 
agglutination of the red blood cells took place on the addition of the 
inactivated serum of the rat, but this did not give rise to any difficulty. A 
further slight increase in the degree of agglutination occurred when human 
blood serum was subsequently added; this did not occur, however, if the 
blood serum of the rabbit was employed as the source of complement, for 
which reason in most of the experiments the latter was used. The amount of 
serum obtained from the rat varied from 0°5 c.c. to 2 cc.; on keeping this 
serum a precipitate of proteid sometimes slowly separated out. 
The above hemolytic system was chosen chiefly because the animals 
employed in its preparation are readily obtainable and can be kept in a 
laboratory. 
As antigen, an alcoholic extract of mouse tumour was prepared by adding 
90 ¢.c. of absolute alcohol to 19 grm. of tumour. The procedure adopted was as 
follows :—The tumour was placed in a mortar, together with the alcohol, and 
was then converted into as fine a pulp as possible. After standing at room 
temperature for six days, with occasional shaking, the alcoholic liquid was 
poured off and filtered. For use in the experiments recorded below, one 
part of the alcoholic extract was added to five parts of 0-85-per-cent. 
solution of sodium chloride. The alcoholic extract was floated on to the top 
of the solution of sodium chloride, and, after standing for about five minutes, 
admixture was completed by shaking the tube, when a very turbid liquid, 
free from precipitate, resulted. 
