GLANDERS 133 
with physiological salt solution and the mixture centrifuged*. The supernatant fluid 
is poured off and the process repeated until all the serum is removed from the corpuscles. 
Enough salt solution is added to the sedimented corpuscles to make the volume of 
blood equal the original amount filtered into the tube.’ 
Hemolytic amboceptor. The hemolytic amboceptor is obtained from the serum 
of a rabbit which has been immunized to the washed red blood corpuscles of a sheep. 
A strong vigorous rabbit is selected and injected with the washed red corpuscles. The 
injections are preferably made intraperitoneally and at intervals of 4-5 days. Two 
ce., 4 ce., 8 cc., and 12 ce. of the corpuscles are respectively used for each injection. 
In seven or eight days after the last injection a small amount of blood is taken from the 
rabbit (by bleeding from an ear vein) and the serum titrated to determine whether its 
hemolytic action is sufficient; that is, whether it will readily dissolve the hemoglobin 
from the corpuscles of a sheep. If it is found satisfactory the animal is bled from the 
carotid and the serum collected and stored in small bottles. It is preferable not to put 
more than 2 cc. in each bottle. The serum may be preserved by adding 0.5 per cent. 
of carbolic acid in a 5 per cent. dilution. If the carbolized serum is not used for three 
days after adding the carbolic acid it will not require inactivation (heating at 56° C. for 
Y% hour). Noguchi prefers adding 2 drops of chloroform to each 2 cc. of hemolytic 
serum and in this case in order to get the best results the serum should be inactivated 
before using. Serum preserved in this way can be kept in the ice-box for from three 
months to a year. It must, however, be titrated at intervals of a few weeks as the 
titre may change. 
Titration of hemolytic rabbit serum. Dilutions of the hemolytic serum are made in 
eight test tubes, according to the following table. It is best to use pipettes to measure 
the required amounts. 
TABLE No. I 
DILUTIONS OF HEMOLYTIC AMBOCEPTOR (RABBIT SERUM). 
Tub NaCl Gi 
Ne a Selution scale : Giuten sce 
1 9 ce. lee. 1-10 | This isa 1-10 basic dilution. 
2 9cc. | 1 cc. of 1-10 1-100 
dilution 
2 8 ce. 2 cc. of 1-100 1-500 
dil. 
4 9 ce. 1 ce. of 1-100 1-1000 
dil. 
5 lee. | 2ec. of 1-1000 1-1500 
dil. 
6 lee. | 1lec.of 1-1000 | 1-2000 
dil. 
7 3 ce. 1 ce. of 1-1000 1-4000 
dil. 
8 8cc. | 2cc. of 1-1000 1-5000 
dil. 
‘Fer hemolytic work 0.85 per cent. (Ehrlich) to 0.9 per cent. (Madsen) salt solution 
is universally employed. We prefer the latter concentration. 
