610 
Jour nal of Agricultural Research 
Vol. XXVIII, No. 7 
A 20-cc. dose has been used in practically all immunizing work that has been 
undertaken with living abortion bacteria, whether under herd conditions, where 
natural Bacterium abortus exposure has prevailed, or in connection with animals 
that have been utilized in a purely experimental capacity. This quantity of 
vaccine represents one-eighth of the growth of abortion organisms developing on 
approximately 15 square inches of 3 per cent glycerin-agar after seven days' 
incubation at a temperature of 37.5° C. When dilutions of the vaccine have 
been plated the number of colonies that have developed has indicated that the 
product contained approximately one billion organisms per cubic centimeter. 
Fifteen to twenty Blake bottles of medium have usually been sown at one time 
with suspensions of as many different strains, and a practice has been made of 
using only such strains as have been artificially cultivated for at least a year. 
After seven days' incubation the growth in each bottle was suspended in 160 cc. 
physiological salt solution. While there has been some variation in the amount of 
growth that has developed in the different bottles, vaccine of fairly uniform 
density has been obtained by combining the suspensions of each lot. The quan¬ 
tity of vaccine administered to each animal has been considerably less than that 
mentioned by Stockman, who refers to the organisms obtained from a similar 
area of medium as constituting a dose for but one or two subjects. 
The vaccinated animals were mated with a bull that was used exclusively for 
serving those so treated and which gave negative complement-fixation and agglu¬ 
tination reactions throughout the experiment. The bull that was mated with 
the bacterin-treated group, as well as those to which the controls were bred, were 
protected from abortion exposure, and gave negative reactions to the serological 
tests at all times. 
The animals receiving bacterin were treated as follows: Two heifers, No. 423 
and 404, received subcutaneous injections two and four times, respectively, 
before being bred, and five times each before being subjected, to Bacterium 
abortus exposure. The injections were continued monthly in doses of 20 cc. 
The suspensions were identical in density with the vaccine described above, but 
were placed in a water bath and maintained at a temperature of 65° C. for one- 
half hour. One heifer, No. 464, was treated bimonthly with half the dose of 
killed organisms and received two injections previously and seven subsequently 
to exposure. The fourth heifer, No. 401, received bimonthly injections of 20 cc., 
two of which wefe given previously to exposure but subsequently to breeding 
date. This animal received seven treatments in all. Individual stalls were 
provided for the treated as well as the control animals during their gestation 
periods, and all the animals were handled in such a manner as to reduce to a 
minimum the possibility of contact with Bacterium abortus infection other than 
that intentionally furnished. 
The method of exposure to Bacterium abortus was as follows: Varying quan¬ 
tities, from 5 to 60 cc., of stomach contents of aborted fetuses, from which 
Bacterium abortus and no other microorganisms could be isolated, were adminis¬ 
tered by the mouth. The material was diluted with from 50 to 100 cc. of physi¬ 
ological salt solution. One feeding was practiced for the most part, although in 
a few instances it was considered desirable to administer the material a second 
time. While numerous lots of stomach contents were required for the exposure 
of the 23 animals utilized, by reason of the fact that the breeding dates extended 
over a period of considerable length, efforts were made to breed the animals at 
such times as would permit of the exposure of those treated and of the controls 
being as nearly uniform in character as possible. 
